刺参（Apostichopus Japonicus Selenka）多糖类免疫增强剂及微生态制剂的研究与应用
|
摘要
本文以我国北方重要的水产养殖经济动物刺参(Apostichopus japonicus Selenka)为研究对象,研究主要的多糖类免疫增强剂(β-葡聚糖、肽聚糖和壳聚糖)对刺参生长、免疫及抗病力的影响。并运用微生物学方法,通过体外实验从刺参肠道、养殖水环境及养殖池底泥中筛选出对刺参有益的潜在益生菌,通过在体养殖实验,研究了筛选出的5株潜在益生菌(TMRC14、TC116、T13、TC22和EN25)对刺参稚参苗(初始平均体重0.2-0.4 g)生长、免疫及抗病力的影响,结果表明菌株T13、TC22和EN25对刺参稚参苗是有益的。因此选取T13、TC22和EN25三株细菌,通过养殖实验,研究在饲料中添加这3株益生菌对刺参幼参(初始平均体重3-5 g)生长、免疫及抗病力的影响,并探讨它们分别与低聚果糖复配的效果。同时对这3株细菌进行了鉴定,并对其最适的培养条件进行了初步摸索。主要研究内容和结果如下:
1.以初始体重为(4.926±0.009) g的刺参为研究对象,探讨了在饲料中添加0 g/kg、1.25 g/kg、2.50 g/kg的β-葡聚糖对刺参生长、免疫及抗病力的影响。实验在循环水系统中进行,每个处理设3个重复。经过8周的养殖实验后,对刺参进行灿烂弧菌(Vibrio splendidus)攻毒。结果表明当饲料中添加1.25 g/kg的β-葡聚糖时,刺参的特定生长率显著高于对照组和2.50 g/kg添加组(P<0.05);后两者之间无显著差异(P>0.05)。饲料中添加β-葡聚糖能显著提高刺参体腔细胞的吞噬活性(P<0.05),而对其呼吸爆发活力的影响并不显著(P>0.05)。随着饲料中β-葡聚糖含量的升高,刺参体腔细胞内酚氧化酶活性显著升高(P<0.05)。刺参体腔细胞内酸性磷酸酶的活性随饲料β-葡聚糖含量升高而有升高的趋势,但各处理组间差异不显著(P>0.05)。攻毒实验表明,1.25 g/kg的β-葡聚糖可以显著降低灿烂弧菌对刺参的致病死亡率(P<0.05)。由此可见,饲料中添加适量的β-葡聚糖(1.25 g/kg)可以提高刺参的特定生长率、非特异性免疫力及抗病能力。
2.以初始体重为(4.931±0.005) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,探讨在基础饲料中分别添加0 mg/kg、200 mg/kg和500 mg/kg的肽聚糖对刺参生长、免疫及抗病力的影响。实验结果表明,随着饲料中肽聚糖含量的升高,刺参的特定生长率呈上升趋势,但各处理之间无显著差异(P>0.05)。饲料中添加肽聚糖显著提高了刺参体腔细胞的吞噬活性(P<0.05),而对其呼吸爆发活力的影响不显著(P>0.05)。饲料中添加200 mg/kg的肽聚糖可以显著提高刺参体腔细胞的酚氧化酶活性(P<0.05),而500 mg/kg添加组与对照组无显著差异(P>0.05)。随着饲料中肽聚糖添加量的升高,刺参体腔细胞内酸性磷酸酶活性呈下降趋势,其中,500 mg/kg肽聚糖添加组显著低于对照组和200 mg/kg的添加组(P<0.05)。攻毒实验表明,饲料中添加肽聚糖可以提高刺参对灿烂弧菌的抵抗能力,其中添加200 mg/kg的肽聚糖显著降低了刺参的致病死亡率(P<0.05)。由此可见,虽然饲料中添加肽聚糖对刺参的生长没有显著影响,但适量的肽聚糖(200 mg/kg)可以显著提高刺参的免疫力和抗病力。
3.以初始体重为(3.734±0.016) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在基础饲料中分别添加0 g/kg、2.5 g/kg、5 g/kg和10 g/kg的壳聚糖,探讨其对刺参生长、免疫及抗病力的影响。实验结果表明,饲料中添加不同梯度的壳聚糖对刺参的生长无显著影响(P>0.05)。当壳聚糖的添加量为10 g/kg时,刺参体腔细胞密度和吞噬活性得到显著提高(P<0.05),但其它添加量组与对照组之间没有显著差异(P>0.05)。饲料中添加壳聚糖对刺参体腔细胞呼吸爆发活力有提高的作用,但影响不显著(P>0.05)。饲料中添加壳聚糖对刺参体腔细胞酚氧化酶活性没有显著影响(P>0.05),但显著降低了刺参体腔细胞内酸性磷酸酶的活力(P<0.05)。攻毒实验表明,饲料中添加壳聚糖并不能显著提高刺参对灿烂弧菌的抵抗能力(P>0.05)。因此,壳聚糖作为免疫增强剂在刺参上的应用需要进一步探讨。
4.用2216E、TSB和MRS培养基,从健康刺参肠道、患病刺参肠道和体壁、刺参养殖池底泥和池水中分离得到细菌182株,以刺参“腐皮综合征”致病菌—灿烂弧菌和假交替单胞菌为指示菌,筛选出对灿烂弧菌或者假交替单胞菌有抑制作用的菌株19株,并对其菌落形态和革兰氏染色情况进行了观察。通过菌株16S rDNA的序列比对,确定19株细菌全部属于芽孢杆菌属。为了进一步筛选刺参潜在的益生菌,对19株细菌的产蛋白酶和淀粉酶性能进行了研究。根据细菌的分离来源及菌落形态和对病原菌的抑制作用情况,选取5株潜在的益生菌用于刺参的安全性实验。结果表明5株潜在的益生菌TMRC14、TC116、T13、TC22和EN25对刺参是安全的,可以成为刺参益生菌的候选菌株,用于后续的实验。
5.以初始体重为(0.204±0.009) g的刺参为研究对象,探讨在基础饲料(鼠尾藻粉)中分别添加10~5、10~7、10~9 CFU/g的TMRC14或者TC116对刺参生长、免疫力及抗病力的影响。养殖实验一共进行30d,静水养殖,每天换水50%。养殖实验结束后,对刺参进行7d灿烂弧菌浸浴攻毒。结果表明,饲料中添加TMRC14对刺参的特定生长率(SGR)没有显著影响(P>0.05);添加10~5-10~7 CFU/g的TC116时,刺参的SGR和对照没有显著区别(P>0.05),但当添加量升高到10~9 CFU/g时,刺参的SGR显著下降(P<0.05)。饲料中添加TMRC14或者TC116对刺参体腔细胞密度没有显著影响(P>0.05)。当TMRC14的添加量为10~9CFU/g时,刺参体腔细胞的吞噬活性显著升高(P<0.05),刺参体腔细胞的呼吸爆发活性也在10~9CFU/g时达到最高,但与对照和其它添加组没有显著差异(P>0.05)。饲料中添加10~7CFU/g TC116时,体腔细胞吞噬活性显著高于对照和其它添加量组(P<0.05)。刺参体腔细胞呼吸爆发活性在TC116的添加量为10~7CFU/g时显著高于对照和其它添加量组(P<0.05),而10~5CFU/g和10~9CFU/g添加组与对照没有显著差异(P>0.05)。攻毒实验表明,饲料中添加10~7CFU/g和10~9CFU/g的TMRC14时,降低了刺参浸浴灿烂弧菌的死亡率,但与对照无显著差异(P>0.05)。饲料中添加10~5CFU/g和10~7CFU/g的TC116时,降低了刺参浸浴灿烂弧菌的死亡率,但与对照并无显著差异(P>0.05)。因此,TMRC14和TC116在刺参生长、免疫和抗病力促进方面的作用并不显著,在刺参稚参苗的生产养殖过程中应用价值不大。
6.以初始体重为(0.204±0.009) g的刺参为研究对象,探讨在基础饲料(鼠尾藻粉)中分别添加10~5 CFU/g、10~7 CFU/g、10~9 CFU/g潜在的益生菌T13或TC22对刺参生长、免疫及抗病力的影响。以初始体重为(0.375±0.024) g的刺参为研究对象,探讨在基础饲料中添加10~5 CFU/g、10~7 CFU/g、10~9 CFU/g潜在的益生菌EN25对刺参生长、免疫及抗病力的影响。养殖实验共进行30d,静水养殖,每天换水50%。养殖实验结束后,对刺参进行7d的灿烂弧菌浸浴攻毒实验。结果表明,当饲料中分别添加10~9 CFU/g的T13或者TC22时,刺参的特定生长率显著升高(P<0.05),而添加EN25对刺参的生长没有显著影响(P>0.05)。饲料中添加T13、TC22、EN25对刺参体腔细胞数量没有显著影响(P>0.05)。饲料中添加10~9 CFU/g的T13或者TC22可以显著提高刺参体腔细胞吞噬活性、呼吸爆发活力和总一氧化氮合酶活力(T-NOS)(P<0.05),但对刺参体腔细胞内超氧化物歧化酶(SOD)、酸性磷酸酶(ACP)活力无显著影响(P>0.05)。当饲料中EN25的添加量为10~7 CFU/g时,刺参的体腔细胞吞噬活性、呼吸爆发活力和T-NOS显著高于对照组(P<0.05),而10~5-10~7 CFU/g的EN25对刺参体腔细胞内SOD、ACP无显著影响(P>0.05);添加量为10~9 CFU/g时,刺参体腔细胞内SOD显著降低(P<0.05),但对ACP无显著影响(P>0.05)。攻毒实验表明,饲料中分别添加10~9 CFU/g的T13或者TC22可以显著降低灿烂弧菌攻毒后刺参的死亡率(P<0.05)。饲料中添加10~7 CFU/g的EN25时,刺参抵御灿烂弧菌感染的能力显著高于对照组(P<0.05)。因此T13、TC22、EN25可以作为益生菌应用到刺参的育苗过程中。
7.以初始体重(4.893±0.028) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在饲料中分别添加0、10~7、10~9 CFU/g的T13,并在每个T13添加水平,分别添加0、0.5%的果糖,以3×2的实验设计配制6种饲料,探讨饲料中添加T13和果糖及两者复配对刺参生长、免疫、肠道菌群及抗病力的影响。养殖实验结束后,每个重复取6头刺参用于免疫指标及肠道菌群分析,剩余刺参用于灿烂弧菌攻毒实验。结果表明,随着饲料中T13含量的升高,刺参SGR有升高的趋势,饲料中单独添加果糖或者与T13复配也提高了刺参的SGR,但所有处理组与对照差异不显著(P>0.05)。饲料中添加T13使刺参体腔细胞吞噬活性、呼吸爆发活性、酚氧化酶活性及一氧化氮合酶活性都得到提高,其中酚氧化酶活性显著高于对照组(P<0.05)。饲料中添加T13显著提高了刺参肠道可培养细菌总数(P<0.05),当T13添加量为10~9 CFU/g时,刺参肠道中弧菌总数显著高于对照(P<0.05),但当添加量为10~7 CFU/g时,刺参肠道弧菌总数与对照没有差异(P>0.05)。饲料中添加10~9 CFU/g T13显著降低了灿烂弧菌攻毒后刺参的死亡率(P<0.05)。饲料中添加0.5%的果糖可以显著提高刺参体腔细胞吞噬活性、呼吸爆发和酚氧化酶活性,并显著降低刺参攻毒后的死亡率(P<0.05)。饲料中果糖与T13复配与否,对刺参肠道可培养细菌总数没有显著影响(P>0.05)。单独添加果糖显著降低了刺参肠道弧菌总数(P<0.05),但果糖与T13复配对刺参肠道弧菌总数影响不显著(P>0.05)。饲料中0.5%果糖与T13复配时,刺参体腔细胞吞噬活性、呼吸爆发、酚氧化酶活性有所升高但和对照没有显著差异(P>0.05),而刺参一氧化氮合酶在果糖与10~9 CFU/g T13复配组显著高于对照组(P<0.05)。果糖与T13复配使刺参抗病力得到显著提高(P<0.05),但与单独添加果糖或者10~9 CFU/g T13相比并没有显著差异(P>0.05)。
8.以初始体重(4.918±0.022)g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在饲料中分别添加0、10~7、10~9 CFU/g的TC22,并在每个TC22的添加梯度,分别添加0、0.5%果糖,以3×2的实验设计配制成6种饲料,探讨饲料中添加TC22和果糖及两者复配对刺参生长、免疫、肠道菌群及抗病力的影响。养殖实验结束后,每个重复取6头刺参用于免疫指标及肠道菌群分析,剩余刺参用于灿烂弧菌攻毒实验。结果表明,在不同的TC22添加梯度下,无论添加果糖与否,刺参的特定生长率都与对照无显著差异(P>0.05)。随着饲料中TC22含量的升高,刺参体腔细胞吞噬活性、呼吸爆发活性及酚氧化酶活性都得到显著提高(P<0.05),而刺参体腔细胞一氧化氮合酶活性呈现先升高后降低的趋势,但与对照组没有显著差异(P>0.05)。饲料中添加10~9 CFU/g TC22显著提高了刺参肠道可培养细菌总数(P<0.05),但刺参肠道中弧菌总数并没有受到饲料中添加TC22和果糖的影响(P>0.05)。饲料中添加10~9 CFU/g TC22显著降低了灿烂弧菌攻毒后刺参的死亡率(P<0.05),添加0.5%的果糖可以显著提高刺参的免疫反应,降低刺参的死亡率(P<0.05)。饲料中0.5%果糖与10~9 CFU/g TC22复配时,刺参的免疫力和抗病力得到显著提高(P<0.05)。因此,益生菌TC22对刺参的健康有促进作用,并且和果糖复配具有正向的交互作用。
9.以初始体重(3.127±0.012) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在饲料中分别添加0、10~7、10~9 CFU/g的EN25,并在每个EN25添加水平,分别添加0、0.5%的果糖,以3×2的实验设计配制成6种饲料,探讨饲料中添加EN25和果糖及两者复配对刺参生长、免疫、肠道菌群及抗病力的影响。养殖实验结束后,每个重复取6头刺参用于免疫指标及肠道菌群分析,剩余刺参用于灿烂弧菌攻毒实验。结果表明,当饲料中添加10~7 CFU/g EN25时,刺参SGR最高,但与对照和其它组差异不显著(P>0.05);饲料中添加果糖或者果糖与EN25复配对刺参SGR无显著影响(P>0.05)。饲料中单独添加EN25时,刺参体腔细胞吞噬活性、呼吸爆发活性显著高于对照组(P<0.05),但其酚氧化酶和一氧化氮合酶活性并没受到显著影响(P>0.05)。与对照相比,饲料中单独添加EN25对刺参肠道可培养细菌总数无显著影响(P>0.05),却显著降低了肠道弧菌总数(P<0.05)。饲料中单独添加EN25显著降低了灿烂弧菌攻毒后刺参的死亡率(P<0.05)。饲料中单独添加0.5%的果糖可以提高刺参体腔细胞吞噬活性、呼吸爆发、酚氧化酶和一氧化氮合酶活性,并显著降低刺参的死亡率(P<0.05)。饲料中0.5%果糖与EN25复配时,刺参体腔细胞吞噬活性、呼吸爆发、酚氧化酶和一氧化氮合酶活性有所升高,其中吞噬活性得到显著提高(P<0.05),呼吸爆发在果糖与10~7 CFU/g EN25复配组显著高于对照组(P<0.05)。饲料中果糖与EN25复配与否,对刺参肠道可培养细菌总数没有显著影响(P>0.05);而当果糖与10~7 CFU/g EN25复配时,刺参肠道弧菌总数显著降低(P<0.05)。果糖与EN25复配提高了刺参抵抗灿烂弧菌的能力,并在EN25的复配浓度为10~9 CFU/g时达到显著水平(P<0.05)。
10~.通过Biolog微生物自动鉴定系统和16S rDNA序列的聚类分析相结合的方法,对筛选出的对刺参育苗和幼体养成都有有益作用的3株益生菌(T13、TC22和EN25)进行了鉴定并对其最适的摇瓶培养条件进行了简单研究,结果表明T13和枯草芽孢杆菌聚为一类,最适培养条件为:接种量2%、装液量50ml/250ml、初始最佳pH 7.0、温度32℃、转速180 r/min。TC22和地衣芽孢杆菌聚为一类,最适培养条件为:接种量4%、装液量75ml/250ml、初始最佳pH 7.2、温度30℃、转速180 r/min。EN25和蜡样芽孢杆菌聚为一类,最适培养条件为:接种量1%、装液量25ml/250ml、初始最佳pH 8.0、温度30℃、转速为180 r/min。
Three feeding experiments were conducted to investigate the effects of three main polysaccharides immunostimulants includingβ-glucan, peptitoglycan and chitosan on growth, immunity and disease resistance in sea cucumber (Apostichopus japonicus Selenka). Nineteen potential probiotics were isolated from intestine of sea cucumber, culturing water and the mud of the pond. Five potential probiotics were selected for further study according to inhibitory ability against phathogen, colony morphologic, species and the source of isolation. Then, the effects of those five potential probiotics on growth, immunity and disease resisitance in juvenile sea cucumber (Initial weight 0.2-0.4 g) were studied. Three strains (T13, TC22 and EN25) of five potential probiotics were found to be beneficial to juvenile sea cucumber. Then three feeding experiments were conducted to investigate the effects of T13, TC22 and EN25 on growth, immunity, microflora and disease resisitance in young sea cucumber (Initial weight 3-5 g). The interaction between three probiotics and fructooligosaccharide respectively was also studied. At the end of the experiment, those three probiotics were identified and the better condition for growth of three probiotics was preliminarily studied. The results are summarized as follows:
1. An 8 week feeding trial was conducted to determine the effects of dietary supplementation ofβ-glucan on the growth, immunity and resistance of sea cucumber against Vibrio splendidus infection. A basal diet was formulated to contain 20.6% crude protein and 4.8% crude lipid. Two levels (1.25 and 2.50 g/kg) ofβ-glucan were added to the basal diet to replace wheat. After the feeding trial, a V. splendidus injection challenge was executed to test the effects ofβ-glucan on disease resistance. Enhanced growth was observed in sea cucumber fed the diet supplemented with 1.25 g glucan/kg, but not in sea cucumber fed the diet supplemented with 2.50 g glucan/kg. The coelomocyte phagocytosis activity of sea cucumber fedβ-glucan supplemented diets was significantly (P<0.05) higher than those of sea cucumbers fed the basal diet. In addition, phenoloxidase activity of coelomocytes was significantly (P<0.05) enhanced by dietary supplementation ofβ-glucan. Sea cucumbers fed 1.25 g/kg glucan had a significant (P<0.05) increase in respiratory burst of coelomocytes, compared to sea cucumber fed diet containing 2.50 gβ-glucan/kg; however, coelomocyte respiratory burst of sea cucumber fed 2.50 gβ-glucan/kg diet was not significantly different from those of sea cucumber fed the basal diet. The challenge test showed that dietary supplementation ofβ-glucan at inclusion level of 1.25 g/kg conferred significant protection to sea cucumber against V. splendidus infection. However, protective effect ofβ-glucan supplementation at 2.50 g/kg was marginal. It is concluded that dietaryβ-glucan has potential for use in diet formulations of sea cucumber to limit the adverse effects of V. splendidus; however, dosage should be an important consideration in administration.
2. An 8 week feeding experiment was conducted to determine the effects of dietary peptidoglycan on growth, immunity and resistance of sea cucumber against Vibrio splendidus infection. The basal diet was supplemented with 0, 200 and 500 mg/kg peptidoglycan to formulate three experimental diets. Results showed that growth of sea cucumber was improved with increasing dietary peptidoglycan. But no significant difference was observed between the control and peptidoglycan supplementation groups. The coelomocyte phagocytosis activity of sea cucumbers fed peptidoglycan supplemented diets was significantly (P<0.05) higher than those of sea cucumbers fed the basal diet. However, respiratory burst of sea cucumber coelomocytes was not significantly influenced by dietary peptidoglycan (P>0.05). Phenoloxidase activity of coelomocytes was significantly increased in animals fed the diet with 200 mg/kg peptidoglycan, but it was not affected by dietary peptidoglycan at 500 mg/kg compared to control (P>0.05). The acid phosphatase activity was decreased with increasing dietary peptidoglycan, and it was significantly lower in sea cucumbers fed the diet with 500 mg/kg peptidoglycan compared to animals fed other two experiment diets (P<0.05). The challenge test showed that dietary peptidoglycan at inclusion level of 200 mg/kg led to significant protection to sea cucumber against V. splendidus infection (P<0.05). It is concluded that dietary peptidoglycan has potential for use in diet formulations of sea cucumber to limit the adverse effects of V. splendidus.
3. 8 week feeding experiment was conducted to determine the effects of dietary chitosan on growth, immunity and resistance of sea cucumber against Vibrio splendidus infection. The basal diet was supplemented with 0, 2.5, 5 and 10~ g/kg chitosan to formulate four experimental diets. Results showed that growth of sea cucumber was not significantly affected by dietary chitosan (P>0.05). The coelomocyte density and coelomocyte phagocytosis activity of sea cucumbers fed chitosan at 10 g/kg was significantly (P<0.05) higher than those of sea cucumbers fed the basal diet. However, respiratory burst of sea cucumber coelomocytes was not significantly influenced by dietary chitosan (P>0.05). There was no significant difference in phenoloxidase activity of coelomocytes (P>0.05). The acid phosphatase activity was significantly decreased in sea cucumbers fed diets with chitosan compared to animals fed the control diet (P<0.05). The challenge test showed that dietary chitosan conferred better protection to sea cucumber against V. splendidus infection, but no significant difference was observed compared to control group (P>0.05). Therefore, further studies on application of chitosan in sea cucumber are necessary.
4. A total of 182 strains were isolated from the intestine of healthy sea cucumbers, intestine and skin of sea cucumbers affected by skin ulcer, mud and sea water of the pond cultured sea cucumbers by 2216E, TSB and MRS bacterial media. All selected isolates were tested for their inhibitory activity to two pathogens of sea cucumber, Vibrio splendidus and Pseudoalteromonas nigrifaciens by spot-on-lawn method. The 19 isolates of those strains were found to be inhibitory to both pathogens or one of them. Morphological analysis was conducted including colony morphologic and Gram coloration. The potential probiotics were identified by 16S rDNA preliminarily. The results showed that all 19 isolates were Gram-positive and Bacillus strains. All strains had amylase activity and 18 of them had proteinase activity. In the end, 5 isolates of 19 strains were selected for the safety test according to source of isolate, colony morphologic and inhibition against pathogens. The safety test of 5 selected potential probiotics showed that these 5 isolates could not induce disease and mortality of sea cucumber. Therefore, these 5 isolates (TMRC14, TC116, T13, TC22 and EN25) could be used for further study to investigate whether these 5 isolates can be applied for sea cucumber to increase the health of animals or not.
5. The effects of potential probiotic TMRC14 and TC116 on growth, immune capacity, and disease resistance in juvenile sea cucumber Apostichopus japonicus were studied. Animals were fed with diet containing TMRC14 or TC116 at 0, 10~5, 10~7 and 10~9 CFU/g for 30 days. At the end of the feeding trial, fifteen sea cucumbers from each aquarium were sampled for immune indices measurement. Then twenty sea cucumbers of every replicate were challenged by immersion with Vibrio splendidus. The results revealed that administration of TMRC14 at all levels or TC116 at 10~5-10~7 CFU/g diet had no significant effect on the growth of sea cucumbers (P>0.05), but the growth of sea cucumber fed with diet containing TC116 at 10~9 CFU/g was significantly decreased (P<0.05). No statistical difference was found in the total coelomocytes counts in sea cucumbers fed the diet containing TMRC14 or TC116 (P>0.05). Phagocytosis of sea cucumber coelomocytes was significantly improved in animals fed with TMRC14 at 10~9 CFU/g diet or TC116 at 10~7 CFU/g diet (P<0.05). Dietary TMRC14 at 10~9 CFU/g increased the respiratory burst activity of sea cucumber coelomocytes, although there was no significant difference (P>0.05). Respiratory burst activity of sea cucumber fed the diet containing TC116 at 10~7 CFU/g was significantly higher than the control and other two groups. The cumulative mortality after V. splendidus challenge was decreased in the sea cucumbers fed with TMRC14 at 10~7-10~9 CFU/g or TC116 at 10~5-10~7 CFU/g, however no significant differences were observed between experimental and the control groups (P>0.05). Therefore, the effect of potential probiotic TMRC14 or TC116 on growth, immunity and protective effect against V. splendidus in sea cucumber were marginal.
6. The effects of potential probiotic T13, TC22 and EN25 on growth, immune capacity, and disease resistance in juvenile sea cucumber Apostichopus japonicus were studied. Animals were fed the diet containing T13, TC22 and EN25 at 0, 10~5, 10~7 and 10~9 CFU/g for 30 days. At the end of the feeding trial, fifteen sea cucumbers from each aquarium were sampled for immune indices measurement. Then twenty sea cucumbers of every replicate were challenged by immersion with Vibrio splendidus. The results revealed that administration of T13 or TC22 at 10~9 CFU/g diet had significant effect on the growth of sea cucumbers (P<0.05), but dietary EN25 had no effect on the growth of animals (P>0.05). Phagocytosis, respiratory burst activity and total nitric oxide synthase (T-NOS) activity of sea cucumber coelomocytes were significantly improved in animals fed the diet with T13 or TC22 at 10~9 CFU/g diet or EN25 at 10~7 CFU/g diet (P<0.05). No statistical difference was found in the total coelomocytes counts and superoxide dismutase (SOD) activity in sea cucumbers fed with diet containing T13 or TC22 at 10~5-10~9 CFU/g or EN25 at 10~5-10~7 CFU/g (P>0.05), however, the SOD activity was significantly decreased in the group fed with diet containing EN25 at 10~9 CFU/g(P<0.05). Dietary supplementation of T13, TC22 or EN25 did not significantly influence acid phosphatase (ACP) activity of sea cucumber coelomocytes. The cumulative mortality after V. splendidus challenge was decreased significantly in the group fed the diet with T13 or TC22 at dose of 10~9 CFU/g feed or EN25 at 10~7 CFU/g feed (P<0.05). The results of present study confirmed the potential beneficial effects of T13, TC22 and EN25 as dietary probiotic in sea cucumber.
7. The effects of probiotic T13 and the synergistic effects of T13 and fructooligosaccharide (FOS) on growth, immune response, microflora and disease resistance in sea cucumber Apostichopus japonicus (initial weight 4.893±0.028 g) were studied. Animals were fed on diet with T13 at doses of 0, 10~7, and 10~9 CFU/g feed with or without 0.5% FOS for 56d. At the end of the feeding trial, six sea cucumbers per tank were sampled for bacterial quantification and immune indices measurement. Then all the sea cucumbers left were challenged by injecting Vibrio splendidus. The results revealed that the SGR of sea cucumber fed the diet containing different doses of T13 with or without FOS was higher than control group, though no significant difference was observed (P>0.05). The phagocytosis activity, respiratory burst activity, phenoloxidase activity and total nitric oxide synthase of sea cucumber fed with diet containing T13 were improved, among these four indices, phenoloxidase activity was significantly higher than control (P<0.05). The total viable bacteria and Vibrio bacteria counts were enhanced significantly in sea cucumber fed with T13 at dose of 10~9 CFU/g feed (P<0.05). The cumulative mortality after V. splendidus challenge was decreased significantly in the group fed with T13 at dose of 10~9 CFU/g feed (P<0.05). The phagocytosis activity, respiratory burst activity and phenoloxidase activity of sea cucumbers fed with 0.5% FOS were increased significantly. And the sea cucumber fed with 0.5% FOS had a significant lower cumulative mortality than the control group (P<0.05). The total viable bacteria counts of sea cucumber were not influenced by dietary FOS with or without T13 (P>0.05). The total Vibrio bacteria counts was significantly decreased in sea cucumber fed with 0.5% FOS (P<0.05), however, it was not affected in sea cucumber fed diet containing FOS and T13 (P>0.05). The combination of 0.5% FOS with T13 increased the phagocytosis activity, respiratory burst activity and phenoloxidase activity of sea cucumbers without significant difference compared to control (P>0.05). The total nitric oxide synthase of sea cucumber fed with diet containing FOS and T13 at 10~9 CFU/g was significantly increased compared to control (P<0.05). The combination of 0.5% FOS with T13 improved disease resistance of sea cucumber compared with control (P<0.05).
8. The effects of probiotic TC22 and the synergistic effects of TC22 and fructooligosaccharide (FOS) on growth, immune capacity, microflora and disease resistance in sea cucumber Apostichopus japonicus (initial weight 4.918±0.022 g) were studied. Animals were fed the diet with TC22 at doses of 0, 10~7, and 10~9 CFU/g feed with or without 0.5% FOS for 56d. At the end of the feeding trial, six sea cucumbers per tank were sampled for bacterial quantification and immune indices measurement. Then all the sea cucumbers left were challenged by injecting Vibrio splendidus. The results revealed that different doses of TC22 with or without FOS had no significant influence on sea cucumber growth (P>0.05). However, along with the increasing of TC22 dosage, the phagocytosis activity, respiratory burst activity and phenoloxidase activity of sea cucumber were improved significantly (P<0.05), while the total nitric oxide synthase acitivity of animals was not significantly influenced (P>0.05). The total viable bacteria counts was significantly enhanced in the sea cucmber fed with TC22 at dose of 10~9 CFU/g feed (P<0.05). But no significant difference was found on total Vibrio counts between trial treatment and the control (P>0.05). The cumulative mortality after V. splendidus challenge decreased significantly in the groups fed with TC22 at dose of 10~9 CFU/g feed (P<0.05). Animals fed with 0.5% FOS showed higher immune response and lower cumulative mortality than the control group (P<0.05). The combination of 0.5% FOS with TC22 at dose of 10~9 CFU/g feed produced significantly positive synergistic effects on sea cucumber immune responses and disease resistance compared with control group(P<0.05). Results of this experiment confirmed the potential of TC22 as dietary probiotic and the synergistic effects of TC22 and FOS in sea cucumber.
9. The effects of probiotic EN25 and the synergistic effects of EN25 and fructooligosaccharide (FOS) on growth, immune response, microflora and disease resistance in sea cucumber Apostichopus japonicus (initial weight 3.127±0.012 g) were studied. Animals were fed the diet with EN25 at doses of 0, 10~7, and 10~9 CFU/g feed with or without 0.5% FOS for 56d. At the end of the feeding trial, six sea cucumbers per tank were sampled for bacterial quantification and immune indices measurement. Then all the sea cucumbers left were challenged by injecting Vibrio splendidus. The results showed that the SGR of sea cucumber was not significantly influenced by dietary EN25 with or without FOS (P>0.05). The phagocytosis activity and respiratory burst was significantly increased by dietary EN25 without FOS (P<0.05). Howerver, the phenoloxidase activity and total nitric oxide synthase of sea cucumber were not significantly influenced by dietary EN25 without FOS (P>0.05). Dietary EN25 without FOS did not produce significant influence on the counts of total viable bacteria of sea cucumber (P>0.05), but it significantly decreased the counts of Vibrio bacteria of animals (P<0.05). The cumulative mortality after V. splendidus challenge was decreased significantly in sea cucumber fed the diet containing EN25 without FOS (P<0.05). Sea cucumbers fed with 0.5% FOS had higher phagocytosis activity, respiratory burst activity, phenoloxidase activity and total nitric oxide synthase than control group, and had a significant lower cumulative mortality after challenged by V. splendidus (P<0.05). The combination of 0.5% FOS and EN25 increased the phagocytosis activity, respiratory burst activity, phenoloxidase activity and total nitric oxide synthase of sea cucumbers. Among these immune indices, the phagocytosis activity was significantly increased in sea cucumber fed FOS and EN25 (P<0.05) and respiratory burst activity was also significantly improved in sea cucumber fed on FOS combined with EN25 at 10~7 CFU/g (P<0.05). The total viable bacteria counts of sea cucumber intestine was not affected by dietary FOS with or without EN25 (P>0.05). The total Vibrio bacteria counts was significantly decreased in sea cucumber fed the diet with 0.5% FOS and EN25 at dose of 10~7 CFU/g (P<0.05). The disease resistance was significantly increased in sea cucumber fed on 0.5% FOS with EN25 at dose of 10~9 CFU/g.
10~. The selected probiotics (T13, TC22 and EN25) were identified by Biolog microbes identification system and the cluster analysis of 16S rDNA. The optimum condition of culture for these three probiotics was preliminarily studied. The results showed that T13 was very similar to Bacillus subtilis and the optimum condition for growth was as follows: inoculation level 2%, liquid volumn 50ml/250ml, initial pH 7.0, temperature 32℃, rotating speed 180 r/min. TC22 was very similar to Bacillus licheniformis and the optimum condition for growth was as follows: inoculation level 4%, liquid volumn 75ml/250ml, initial pH 7.2, temperature 30℃, rotating speed 180 r/min. EN25 was very similar to Bacillus cereus and the optimum condition for growth was as follows: inoculation level 1%, liquid volumn 25ml/250ml, initial pH 8.0, temperature 30℃, rotating speed 180 r/min.
1.以初始体重为(4.926±0.009) g的刺参为研究对象,探讨了在饲料中添加0 g/kg、1.25 g/kg、2.50 g/kg的β-葡聚糖对刺参生长、免疫及抗病力的影响。实验在循环水系统中进行,每个处理设3个重复。经过8周的养殖实验后,对刺参进行灿烂弧菌(Vibrio splendidus)攻毒。结果表明当饲料中添加1.25 g/kg的β-葡聚糖时,刺参的特定生长率显著高于对照组和2.50 g/kg添加组(P<0.05);后两者之间无显著差异(P>0.05)。饲料中添加β-葡聚糖能显著提高刺参体腔细胞的吞噬活性(P<0.05),而对其呼吸爆发活力的影响并不显著(P>0.05)。随着饲料中β-葡聚糖含量的升高,刺参体腔细胞内酚氧化酶活性显著升高(P<0.05)。刺参体腔细胞内酸性磷酸酶的活性随饲料β-葡聚糖含量升高而有升高的趋势,但各处理组间差异不显著(P>0.05)。攻毒实验表明,1.25 g/kg的β-葡聚糖可以显著降低灿烂弧菌对刺参的致病死亡率(P<0.05)。由此可见,饲料中添加适量的β-葡聚糖(1.25 g/kg)可以提高刺参的特定生长率、非特异性免疫力及抗病能力。
2.以初始体重为(4.931±0.005) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,探讨在基础饲料中分别添加0 mg/kg、200 mg/kg和500 mg/kg的肽聚糖对刺参生长、免疫及抗病力的影响。实验结果表明,随着饲料中肽聚糖含量的升高,刺参的特定生长率呈上升趋势,但各处理之间无显著差异(P>0.05)。饲料中添加肽聚糖显著提高了刺参体腔细胞的吞噬活性(P<0.05),而对其呼吸爆发活力的影响不显著(P>0.05)。饲料中添加200 mg/kg的肽聚糖可以显著提高刺参体腔细胞的酚氧化酶活性(P<0.05),而500 mg/kg添加组与对照组无显著差异(P>0.05)。随着饲料中肽聚糖添加量的升高,刺参体腔细胞内酸性磷酸酶活性呈下降趋势,其中,500 mg/kg肽聚糖添加组显著低于对照组和200 mg/kg的添加组(P<0.05)。攻毒实验表明,饲料中添加肽聚糖可以提高刺参对灿烂弧菌的抵抗能力,其中添加200 mg/kg的肽聚糖显著降低了刺参的致病死亡率(P<0.05)。由此可见,虽然饲料中添加肽聚糖对刺参的生长没有显著影响,但适量的肽聚糖(200 mg/kg)可以显著提高刺参的免疫力和抗病力。
3.以初始体重为(3.734±0.016) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在基础饲料中分别添加0 g/kg、2.5 g/kg、5 g/kg和10 g/kg的壳聚糖,探讨其对刺参生长、免疫及抗病力的影响。实验结果表明,饲料中添加不同梯度的壳聚糖对刺参的生长无显著影响(P>0.05)。当壳聚糖的添加量为10 g/kg时,刺参体腔细胞密度和吞噬活性得到显著提高(P<0.05),但其它添加量组与对照组之间没有显著差异(P>0.05)。饲料中添加壳聚糖对刺参体腔细胞呼吸爆发活力有提高的作用,但影响不显著(P>0.05)。饲料中添加壳聚糖对刺参体腔细胞酚氧化酶活性没有显著影响(P>0.05),但显著降低了刺参体腔细胞内酸性磷酸酶的活力(P<0.05)。攻毒实验表明,饲料中添加壳聚糖并不能显著提高刺参对灿烂弧菌的抵抗能力(P>0.05)。因此,壳聚糖作为免疫增强剂在刺参上的应用需要进一步探讨。
4.用2216E、TSB和MRS培养基,从健康刺参肠道、患病刺参肠道和体壁、刺参养殖池底泥和池水中分离得到细菌182株,以刺参“腐皮综合征”致病菌—灿烂弧菌和假交替单胞菌为指示菌,筛选出对灿烂弧菌或者假交替单胞菌有抑制作用的菌株19株,并对其菌落形态和革兰氏染色情况进行了观察。通过菌株16S rDNA的序列比对,确定19株细菌全部属于芽孢杆菌属。为了进一步筛选刺参潜在的益生菌,对19株细菌的产蛋白酶和淀粉酶性能进行了研究。根据细菌的分离来源及菌落形态和对病原菌的抑制作用情况,选取5株潜在的益生菌用于刺参的安全性实验。结果表明5株潜在的益生菌TMRC14、TC116、T13、TC22和EN25对刺参是安全的,可以成为刺参益生菌的候选菌株,用于后续的实验。
5.以初始体重为(0.204±0.009) g的刺参为研究对象,探讨在基础饲料(鼠尾藻粉)中分别添加10~5、10~7、10~9 CFU/g的TMRC14或者TC116对刺参生长、免疫力及抗病力的影响。养殖实验一共进行30d,静水养殖,每天换水50%。养殖实验结束后,对刺参进行7d灿烂弧菌浸浴攻毒。结果表明,饲料中添加TMRC14对刺参的特定生长率(SGR)没有显著影响(P>0.05);添加10~5-10~7 CFU/g的TC116时,刺参的SGR和对照没有显著区别(P>0.05),但当添加量升高到10~9 CFU/g时,刺参的SGR显著下降(P<0.05)。饲料中添加TMRC14或者TC116对刺参体腔细胞密度没有显著影响(P>0.05)。当TMRC14的添加量为10~9CFU/g时,刺参体腔细胞的吞噬活性显著升高(P<0.05),刺参体腔细胞的呼吸爆发活性也在10~9CFU/g时达到最高,但与对照和其它添加组没有显著差异(P>0.05)。饲料中添加10~7CFU/g TC116时,体腔细胞吞噬活性显著高于对照和其它添加量组(P<0.05)。刺参体腔细胞呼吸爆发活性在TC116的添加量为10~7CFU/g时显著高于对照和其它添加量组(P<0.05),而10~5CFU/g和10~9CFU/g添加组与对照没有显著差异(P>0.05)。攻毒实验表明,饲料中添加10~7CFU/g和10~9CFU/g的TMRC14时,降低了刺参浸浴灿烂弧菌的死亡率,但与对照无显著差异(P>0.05)。饲料中添加10~5CFU/g和10~7CFU/g的TC116时,降低了刺参浸浴灿烂弧菌的死亡率,但与对照并无显著差异(P>0.05)。因此,TMRC14和TC116在刺参生长、免疫和抗病力促进方面的作用并不显著,在刺参稚参苗的生产养殖过程中应用价值不大。
6.以初始体重为(0.204±0.009) g的刺参为研究对象,探讨在基础饲料(鼠尾藻粉)中分别添加10~5 CFU/g、10~7 CFU/g、10~9 CFU/g潜在的益生菌T13或TC22对刺参生长、免疫及抗病力的影响。以初始体重为(0.375±0.024) g的刺参为研究对象,探讨在基础饲料中添加10~5 CFU/g、10~7 CFU/g、10~9 CFU/g潜在的益生菌EN25对刺参生长、免疫及抗病力的影响。养殖实验共进行30d,静水养殖,每天换水50%。养殖实验结束后,对刺参进行7d的灿烂弧菌浸浴攻毒实验。结果表明,当饲料中分别添加10~9 CFU/g的T13或者TC22时,刺参的特定生长率显著升高(P<0.05),而添加EN25对刺参的生长没有显著影响(P>0.05)。饲料中添加T13、TC22、EN25对刺参体腔细胞数量没有显著影响(P>0.05)。饲料中添加10~9 CFU/g的T13或者TC22可以显著提高刺参体腔细胞吞噬活性、呼吸爆发活力和总一氧化氮合酶活力(T-NOS)(P<0.05),但对刺参体腔细胞内超氧化物歧化酶(SOD)、酸性磷酸酶(ACP)活力无显著影响(P>0.05)。当饲料中EN25的添加量为10~7 CFU/g时,刺参的体腔细胞吞噬活性、呼吸爆发活力和T-NOS显著高于对照组(P<0.05),而10~5-10~7 CFU/g的EN25对刺参体腔细胞内SOD、ACP无显著影响(P>0.05);添加量为10~9 CFU/g时,刺参体腔细胞内SOD显著降低(P<0.05),但对ACP无显著影响(P>0.05)。攻毒实验表明,饲料中分别添加10~9 CFU/g的T13或者TC22可以显著降低灿烂弧菌攻毒后刺参的死亡率(P<0.05)。饲料中添加10~7 CFU/g的EN25时,刺参抵御灿烂弧菌感染的能力显著高于对照组(P<0.05)。因此T13、TC22、EN25可以作为益生菌应用到刺参的育苗过程中。
7.以初始体重(4.893±0.028) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在饲料中分别添加0、10~7、10~9 CFU/g的T13,并在每个T13添加水平,分别添加0、0.5%的果糖,以3×2的实验设计配制6种饲料,探讨饲料中添加T13和果糖及两者复配对刺参生长、免疫、肠道菌群及抗病力的影响。养殖实验结束后,每个重复取6头刺参用于免疫指标及肠道菌群分析,剩余刺参用于灿烂弧菌攻毒实验。结果表明,随着饲料中T13含量的升高,刺参SGR有升高的趋势,饲料中单独添加果糖或者与T13复配也提高了刺参的SGR,但所有处理组与对照差异不显著(P>0.05)。饲料中添加T13使刺参体腔细胞吞噬活性、呼吸爆发活性、酚氧化酶活性及一氧化氮合酶活性都得到提高,其中酚氧化酶活性显著高于对照组(P<0.05)。饲料中添加T13显著提高了刺参肠道可培养细菌总数(P<0.05),当T13添加量为10~9 CFU/g时,刺参肠道中弧菌总数显著高于对照(P<0.05),但当添加量为10~7 CFU/g时,刺参肠道弧菌总数与对照没有差异(P>0.05)。饲料中添加10~9 CFU/g T13显著降低了灿烂弧菌攻毒后刺参的死亡率(P<0.05)。饲料中添加0.5%的果糖可以显著提高刺参体腔细胞吞噬活性、呼吸爆发和酚氧化酶活性,并显著降低刺参攻毒后的死亡率(P<0.05)。饲料中果糖与T13复配与否,对刺参肠道可培养细菌总数没有显著影响(P>0.05)。单独添加果糖显著降低了刺参肠道弧菌总数(P<0.05),但果糖与T13复配对刺参肠道弧菌总数影响不显著(P>0.05)。饲料中0.5%果糖与T13复配时,刺参体腔细胞吞噬活性、呼吸爆发、酚氧化酶活性有所升高但和对照没有显著差异(P>0.05),而刺参一氧化氮合酶在果糖与10~9 CFU/g T13复配组显著高于对照组(P<0.05)。果糖与T13复配使刺参抗病力得到显著提高(P<0.05),但与单独添加果糖或者10~9 CFU/g T13相比并没有显著差异(P>0.05)。
8.以初始体重(4.918±0.022)g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在饲料中分别添加0、10~7、10~9 CFU/g的TC22,并在每个TC22的添加梯度,分别添加0、0.5%果糖,以3×2的实验设计配制成6种饲料,探讨饲料中添加TC22和果糖及两者复配对刺参生长、免疫、肠道菌群及抗病力的影响。养殖实验结束后,每个重复取6头刺参用于免疫指标及肠道菌群分析,剩余刺参用于灿烂弧菌攻毒实验。结果表明,在不同的TC22添加梯度下,无论添加果糖与否,刺参的特定生长率都与对照无显著差异(P>0.05)。随着饲料中TC22含量的升高,刺参体腔细胞吞噬活性、呼吸爆发活性及酚氧化酶活性都得到显著提高(P<0.05),而刺参体腔细胞一氧化氮合酶活性呈现先升高后降低的趋势,但与对照组没有显著差异(P>0.05)。饲料中添加10~9 CFU/g TC22显著提高了刺参肠道可培养细菌总数(P<0.05),但刺参肠道中弧菌总数并没有受到饲料中添加TC22和果糖的影响(P>0.05)。饲料中添加10~9 CFU/g TC22显著降低了灿烂弧菌攻毒后刺参的死亡率(P<0.05),添加0.5%的果糖可以显著提高刺参的免疫反应,降低刺参的死亡率(P<0.05)。饲料中0.5%果糖与10~9 CFU/g TC22复配时,刺参的免疫力和抗病力得到显著提高(P<0.05)。因此,益生菌TC22对刺参的健康有促进作用,并且和果糖复配具有正向的交互作用。
9.以初始体重(3.127±0.012) g的刺参为研究对象,在循环水系统中进行8周的养殖实验,在饲料中分别添加0、10~7、10~9 CFU/g的EN25,并在每个EN25添加水平,分别添加0、0.5%的果糖,以3×2的实验设计配制成6种饲料,探讨饲料中添加EN25和果糖及两者复配对刺参生长、免疫、肠道菌群及抗病力的影响。养殖实验结束后,每个重复取6头刺参用于免疫指标及肠道菌群分析,剩余刺参用于灿烂弧菌攻毒实验。结果表明,当饲料中添加10~7 CFU/g EN25时,刺参SGR最高,但与对照和其它组差异不显著(P>0.05);饲料中添加果糖或者果糖与EN25复配对刺参SGR无显著影响(P>0.05)。饲料中单独添加EN25时,刺参体腔细胞吞噬活性、呼吸爆发活性显著高于对照组(P<0.05),但其酚氧化酶和一氧化氮合酶活性并没受到显著影响(P>0.05)。与对照相比,饲料中单独添加EN25对刺参肠道可培养细菌总数无显著影响(P>0.05),却显著降低了肠道弧菌总数(P<0.05)。饲料中单独添加EN25显著降低了灿烂弧菌攻毒后刺参的死亡率(P<0.05)。饲料中单独添加0.5%的果糖可以提高刺参体腔细胞吞噬活性、呼吸爆发、酚氧化酶和一氧化氮合酶活性,并显著降低刺参的死亡率(P<0.05)。饲料中0.5%果糖与EN25复配时,刺参体腔细胞吞噬活性、呼吸爆发、酚氧化酶和一氧化氮合酶活性有所升高,其中吞噬活性得到显著提高(P<0.05),呼吸爆发在果糖与10~7 CFU/g EN25复配组显著高于对照组(P<0.05)。饲料中果糖与EN25复配与否,对刺参肠道可培养细菌总数没有显著影响(P>0.05);而当果糖与10~7 CFU/g EN25复配时,刺参肠道弧菌总数显著降低(P<0.05)。果糖与EN25复配提高了刺参抵抗灿烂弧菌的能力,并在EN25的复配浓度为10~9 CFU/g时达到显著水平(P<0.05)。
10~.通过Biolog微生物自动鉴定系统和16S rDNA序列的聚类分析相结合的方法,对筛选出的对刺参育苗和幼体养成都有有益作用的3株益生菌(T13、TC22和EN25)进行了鉴定并对其最适的摇瓶培养条件进行了简单研究,结果表明T13和枯草芽孢杆菌聚为一类,最适培养条件为:接种量2%、装液量50ml/250ml、初始最佳pH 7.0、温度32℃、转速180 r/min。TC22和地衣芽孢杆菌聚为一类,最适培养条件为:接种量4%、装液量75ml/250ml、初始最佳pH 7.2、温度30℃、转速180 r/min。EN25和蜡样芽孢杆菌聚为一类,最适培养条件为:接种量1%、装液量25ml/250ml、初始最佳pH 8.0、温度30℃、转速为180 r/min。
Three feeding experiments were conducted to investigate the effects of three main polysaccharides immunostimulants includingβ-glucan, peptitoglycan and chitosan on growth, immunity and disease resistance in sea cucumber (Apostichopus japonicus Selenka). Nineteen potential probiotics were isolated from intestine of sea cucumber, culturing water and the mud of the pond. Five potential probiotics were selected for further study according to inhibitory ability against phathogen, colony morphologic, species and the source of isolation. Then, the effects of those five potential probiotics on growth, immunity and disease resisitance in juvenile sea cucumber (Initial weight 0.2-0.4 g) were studied. Three strains (T13, TC22 and EN25) of five potential probiotics were found to be beneficial to juvenile sea cucumber. Then three feeding experiments were conducted to investigate the effects of T13, TC22 and EN25 on growth, immunity, microflora and disease resisitance in young sea cucumber (Initial weight 3-5 g). The interaction between three probiotics and fructooligosaccharide respectively was also studied. At the end of the experiment, those three probiotics were identified and the better condition for growth of three probiotics was preliminarily studied. The results are summarized as follows:
1. An 8 week feeding trial was conducted to determine the effects of dietary supplementation ofβ-glucan on the growth, immunity and resistance of sea cucumber against Vibrio splendidus infection. A basal diet was formulated to contain 20.6% crude protein and 4.8% crude lipid. Two levels (1.25 and 2.50 g/kg) ofβ-glucan were added to the basal diet to replace wheat. After the feeding trial, a V. splendidus injection challenge was executed to test the effects ofβ-glucan on disease resistance. Enhanced growth was observed in sea cucumber fed the diet supplemented with 1.25 g glucan/kg, but not in sea cucumber fed the diet supplemented with 2.50 g glucan/kg. The coelomocyte phagocytosis activity of sea cucumber fedβ-glucan supplemented diets was significantly (P<0.05) higher than those of sea cucumbers fed the basal diet. In addition, phenoloxidase activity of coelomocytes was significantly (P<0.05) enhanced by dietary supplementation ofβ-glucan. Sea cucumbers fed 1.25 g/kg glucan had a significant (P<0.05) increase in respiratory burst of coelomocytes, compared to sea cucumber fed diet containing 2.50 gβ-glucan/kg; however, coelomocyte respiratory burst of sea cucumber fed 2.50 gβ-glucan/kg diet was not significantly different from those of sea cucumber fed the basal diet. The challenge test showed that dietary supplementation ofβ-glucan at inclusion level of 1.25 g/kg conferred significant protection to sea cucumber against V. splendidus infection. However, protective effect ofβ-glucan supplementation at 2.50 g/kg was marginal. It is concluded that dietaryβ-glucan has potential for use in diet formulations of sea cucumber to limit the adverse effects of V. splendidus; however, dosage should be an important consideration in administration.
2. An 8 week feeding experiment was conducted to determine the effects of dietary peptidoglycan on growth, immunity and resistance of sea cucumber against Vibrio splendidus infection. The basal diet was supplemented with 0, 200 and 500 mg/kg peptidoglycan to formulate three experimental diets. Results showed that growth of sea cucumber was improved with increasing dietary peptidoglycan. But no significant difference was observed between the control and peptidoglycan supplementation groups. The coelomocyte phagocytosis activity of sea cucumbers fed peptidoglycan supplemented diets was significantly (P<0.05) higher than those of sea cucumbers fed the basal diet. However, respiratory burst of sea cucumber coelomocytes was not significantly influenced by dietary peptidoglycan (P>0.05). Phenoloxidase activity of coelomocytes was significantly increased in animals fed the diet with 200 mg/kg peptidoglycan, but it was not affected by dietary peptidoglycan at 500 mg/kg compared to control (P>0.05). The acid phosphatase activity was decreased with increasing dietary peptidoglycan, and it was significantly lower in sea cucumbers fed the diet with 500 mg/kg peptidoglycan compared to animals fed other two experiment diets (P<0.05). The challenge test showed that dietary peptidoglycan at inclusion level of 200 mg/kg led to significant protection to sea cucumber against V. splendidus infection (P<0.05). It is concluded that dietary peptidoglycan has potential for use in diet formulations of sea cucumber to limit the adverse effects of V. splendidus.
3. 8 week feeding experiment was conducted to determine the effects of dietary chitosan on growth, immunity and resistance of sea cucumber against Vibrio splendidus infection. The basal diet was supplemented with 0, 2.5, 5 and 10~ g/kg chitosan to formulate four experimental diets. Results showed that growth of sea cucumber was not significantly affected by dietary chitosan (P>0.05). The coelomocyte density and coelomocyte phagocytosis activity of sea cucumbers fed chitosan at 10 g/kg was significantly (P<0.05) higher than those of sea cucumbers fed the basal diet. However, respiratory burst of sea cucumber coelomocytes was not significantly influenced by dietary chitosan (P>0.05). There was no significant difference in phenoloxidase activity of coelomocytes (P>0.05). The acid phosphatase activity was significantly decreased in sea cucumbers fed diets with chitosan compared to animals fed the control diet (P<0.05). The challenge test showed that dietary chitosan conferred better protection to sea cucumber against V. splendidus infection, but no significant difference was observed compared to control group (P>0.05). Therefore, further studies on application of chitosan in sea cucumber are necessary.
4. A total of 182 strains were isolated from the intestine of healthy sea cucumbers, intestine and skin of sea cucumbers affected by skin ulcer, mud and sea water of the pond cultured sea cucumbers by 2216E, TSB and MRS bacterial media. All selected isolates were tested for their inhibitory activity to two pathogens of sea cucumber, Vibrio splendidus and Pseudoalteromonas nigrifaciens by spot-on-lawn method. The 19 isolates of those strains were found to be inhibitory to both pathogens or one of them. Morphological analysis was conducted including colony morphologic and Gram coloration. The potential probiotics were identified by 16S rDNA preliminarily. The results showed that all 19 isolates were Gram-positive and Bacillus strains. All strains had amylase activity and 18 of them had proteinase activity. In the end, 5 isolates of 19 strains were selected for the safety test according to source of isolate, colony morphologic and inhibition against pathogens. The safety test of 5 selected potential probiotics showed that these 5 isolates could not induce disease and mortality of sea cucumber. Therefore, these 5 isolates (TMRC14, TC116, T13, TC22 and EN25) could be used for further study to investigate whether these 5 isolates can be applied for sea cucumber to increase the health of animals or not.
5. The effects of potential probiotic TMRC14 and TC116 on growth, immune capacity, and disease resistance in juvenile sea cucumber Apostichopus japonicus were studied. Animals were fed with diet containing TMRC14 or TC116 at 0, 10~5, 10~7 and 10~9 CFU/g for 30 days. At the end of the feeding trial, fifteen sea cucumbers from each aquarium were sampled for immune indices measurement. Then twenty sea cucumbers of every replicate were challenged by immersion with Vibrio splendidus. The results revealed that administration of TMRC14 at all levels or TC116 at 10~5-10~7 CFU/g diet had no significant effect on the growth of sea cucumbers (P>0.05), but the growth of sea cucumber fed with diet containing TC116 at 10~9 CFU/g was significantly decreased (P<0.05). No statistical difference was found in the total coelomocytes counts in sea cucumbers fed the diet containing TMRC14 or TC116 (P>0.05). Phagocytosis of sea cucumber coelomocytes was significantly improved in animals fed with TMRC14 at 10~9 CFU/g diet or TC116 at 10~7 CFU/g diet (P<0.05). Dietary TMRC14 at 10~9 CFU/g increased the respiratory burst activity of sea cucumber coelomocytes, although there was no significant difference (P>0.05). Respiratory burst activity of sea cucumber fed the diet containing TC116 at 10~7 CFU/g was significantly higher than the control and other two groups. The cumulative mortality after V. splendidus challenge was decreased in the sea cucumbers fed with TMRC14 at 10~7-10~9 CFU/g or TC116 at 10~5-10~7 CFU/g, however no significant differences were observed between experimental and the control groups (P>0.05). Therefore, the effect of potential probiotic TMRC14 or TC116 on growth, immunity and protective effect against V. splendidus in sea cucumber were marginal.
6. The effects of potential probiotic T13, TC22 and EN25 on growth, immune capacity, and disease resistance in juvenile sea cucumber Apostichopus japonicus were studied. Animals were fed the diet containing T13, TC22 and EN25 at 0, 10~5, 10~7 and 10~9 CFU/g for 30 days. At the end of the feeding trial, fifteen sea cucumbers from each aquarium were sampled for immune indices measurement. Then twenty sea cucumbers of every replicate were challenged by immersion with Vibrio splendidus. The results revealed that administration of T13 or TC22 at 10~9 CFU/g diet had significant effect on the growth of sea cucumbers (P<0.05), but dietary EN25 had no effect on the growth of animals (P>0.05). Phagocytosis, respiratory burst activity and total nitric oxide synthase (T-NOS) activity of sea cucumber coelomocytes were significantly improved in animals fed the diet with T13 or TC22 at 10~9 CFU/g diet or EN25 at 10~7 CFU/g diet (P<0.05). No statistical difference was found in the total coelomocytes counts and superoxide dismutase (SOD) activity in sea cucumbers fed with diet containing T13 or TC22 at 10~5-10~9 CFU/g or EN25 at 10~5-10~7 CFU/g (P>0.05), however, the SOD activity was significantly decreased in the group fed with diet containing EN25 at 10~9 CFU/g(P<0.05). Dietary supplementation of T13, TC22 or EN25 did not significantly influence acid phosphatase (ACP) activity of sea cucumber coelomocytes. The cumulative mortality after V. splendidus challenge was decreased significantly in the group fed the diet with T13 or TC22 at dose of 10~9 CFU/g feed or EN25 at 10~7 CFU/g feed (P<0.05). The results of present study confirmed the potential beneficial effects of T13, TC22 and EN25 as dietary probiotic in sea cucumber.
7. The effects of probiotic T13 and the synergistic effects of T13 and fructooligosaccharide (FOS) on growth, immune response, microflora and disease resistance in sea cucumber Apostichopus japonicus (initial weight 4.893±0.028 g) were studied. Animals were fed on diet with T13 at doses of 0, 10~7, and 10~9 CFU/g feed with or without 0.5% FOS for 56d. At the end of the feeding trial, six sea cucumbers per tank were sampled for bacterial quantification and immune indices measurement. Then all the sea cucumbers left were challenged by injecting Vibrio splendidus. The results revealed that the SGR of sea cucumber fed the diet containing different doses of T13 with or without FOS was higher than control group, though no significant difference was observed (P>0.05). The phagocytosis activity, respiratory burst activity, phenoloxidase activity and total nitric oxide synthase of sea cucumber fed with diet containing T13 were improved, among these four indices, phenoloxidase activity was significantly higher than control (P<0.05). The total viable bacteria and Vibrio bacteria counts were enhanced significantly in sea cucumber fed with T13 at dose of 10~9 CFU/g feed (P<0.05). The cumulative mortality after V. splendidus challenge was decreased significantly in the group fed with T13 at dose of 10~9 CFU/g feed (P<0.05). The phagocytosis activity, respiratory burst activity and phenoloxidase activity of sea cucumbers fed with 0.5% FOS were increased significantly. And the sea cucumber fed with 0.5% FOS had a significant lower cumulative mortality than the control group (P<0.05). The total viable bacteria counts of sea cucumber were not influenced by dietary FOS with or without T13 (P>0.05). The total Vibrio bacteria counts was significantly decreased in sea cucumber fed with 0.5% FOS (P<0.05), however, it was not affected in sea cucumber fed diet containing FOS and T13 (P>0.05). The combination of 0.5% FOS with T13 increased the phagocytosis activity, respiratory burst activity and phenoloxidase activity of sea cucumbers without significant difference compared to control (P>0.05). The total nitric oxide synthase of sea cucumber fed with diet containing FOS and T13 at 10~9 CFU/g was significantly increased compared to control (P<0.05). The combination of 0.5% FOS with T13 improved disease resistance of sea cucumber compared with control (P<0.05).
8. The effects of probiotic TC22 and the synergistic effects of TC22 and fructooligosaccharide (FOS) on growth, immune capacity, microflora and disease resistance in sea cucumber Apostichopus japonicus (initial weight 4.918±0.022 g) were studied. Animals were fed the diet with TC22 at doses of 0, 10~7, and 10~9 CFU/g feed with or without 0.5% FOS for 56d. At the end of the feeding trial, six sea cucumbers per tank were sampled for bacterial quantification and immune indices measurement. Then all the sea cucumbers left were challenged by injecting Vibrio splendidus. The results revealed that different doses of TC22 with or without FOS had no significant influence on sea cucumber growth (P>0.05). However, along with the increasing of TC22 dosage, the phagocytosis activity, respiratory burst activity and phenoloxidase activity of sea cucumber were improved significantly (P<0.05), while the total nitric oxide synthase acitivity of animals was not significantly influenced (P>0.05). The total viable bacteria counts was significantly enhanced in the sea cucmber fed with TC22 at dose of 10~9 CFU/g feed (P<0.05). But no significant difference was found on total Vibrio counts between trial treatment and the control (P>0.05). The cumulative mortality after V. splendidus challenge decreased significantly in the groups fed with TC22 at dose of 10~9 CFU/g feed (P<0.05). Animals fed with 0.5% FOS showed higher immune response and lower cumulative mortality than the control group (P<0.05). The combination of 0.5% FOS with TC22 at dose of 10~9 CFU/g feed produced significantly positive synergistic effects on sea cucumber immune responses and disease resistance compared with control group(P<0.05). Results of this experiment confirmed the potential of TC22 as dietary probiotic and the synergistic effects of TC22 and FOS in sea cucumber.
9. The effects of probiotic EN25 and the synergistic effects of EN25 and fructooligosaccharide (FOS) on growth, immune response, microflora and disease resistance in sea cucumber Apostichopus japonicus (initial weight 3.127±0.012 g) were studied. Animals were fed the diet with EN25 at doses of 0, 10~7, and 10~9 CFU/g feed with or without 0.5% FOS for 56d. At the end of the feeding trial, six sea cucumbers per tank were sampled for bacterial quantification and immune indices measurement. Then all the sea cucumbers left were challenged by injecting Vibrio splendidus. The results showed that the SGR of sea cucumber was not significantly influenced by dietary EN25 with or without FOS (P>0.05). The phagocytosis activity and respiratory burst was significantly increased by dietary EN25 without FOS (P<0.05). Howerver, the phenoloxidase activity and total nitric oxide synthase of sea cucumber were not significantly influenced by dietary EN25 without FOS (P>0.05). Dietary EN25 without FOS did not produce significant influence on the counts of total viable bacteria of sea cucumber (P>0.05), but it significantly decreased the counts of Vibrio bacteria of animals (P<0.05). The cumulative mortality after V. splendidus challenge was decreased significantly in sea cucumber fed the diet containing EN25 without FOS (P<0.05). Sea cucumbers fed with 0.5% FOS had higher phagocytosis activity, respiratory burst activity, phenoloxidase activity and total nitric oxide synthase than control group, and had a significant lower cumulative mortality after challenged by V. splendidus (P<0.05). The combination of 0.5% FOS and EN25 increased the phagocytosis activity, respiratory burst activity, phenoloxidase activity and total nitric oxide synthase of sea cucumbers. Among these immune indices, the phagocytosis activity was significantly increased in sea cucumber fed FOS and EN25 (P<0.05) and respiratory burst activity was also significantly improved in sea cucumber fed on FOS combined with EN25 at 10~7 CFU/g (P<0.05). The total viable bacteria counts of sea cucumber intestine was not affected by dietary FOS with or without EN25 (P>0.05). The total Vibrio bacteria counts was significantly decreased in sea cucumber fed the diet with 0.5% FOS and EN25 at dose of 10~7 CFU/g (P<0.05). The disease resistance was significantly increased in sea cucumber fed on 0.5% FOS with EN25 at dose of 10~9 CFU/g.
10~. The selected probiotics (T13, TC22 and EN25) were identified by Biolog microbes identification system and the cluster analysis of 16S rDNA. The optimum condition of culture for these three probiotics was preliminarily studied. The results showed that T13 was very similar to Bacillus subtilis and the optimum condition for growth was as follows: inoculation level 2%, liquid volumn 50ml/250ml, initial pH 7.0, temperature 32℃, rotating speed 180 r/min. TC22 was very similar to Bacillus licheniformis and the optimum condition for growth was as follows: inoculation level 4%, liquid volumn 75ml/250ml, initial pH 7.2, temperature 30℃, rotating speed 180 r/min. EN25 was very similar to Bacillus cereus and the optimum condition for growth was as follows: inoculation level 1%, liquid volumn 25ml/250ml, initial pH 8.0, temperature 30℃, rotating speed 180 r/min.
引文
Ai Q H, Mai K S, Zhang L, et al. Effects of dietaryβ-1,3-glucan on innate immune response of large yellow croaker, Pseudosciaena crocea. Fish Shellfish Immunol. 2007, 22: 394-402
Ainsworth A J. A beta-glucan inhibitable zymosan receptor on channel catfish neutrophils. Vet. Immunol Immunopathol., 1994, 41: 141-152
Aly S M, Abd-El-Rahman A M, John G, et al. Characterization of Some Bacteria Isolated from Oreochromis niloticus and their Potential Use as Probiotics. Aquaculture, 2008a, 277: 1-6
Aly S M, Ahmed Y A, Ghareeb A A, et al. Studies on Bacillus subtilis and Lactobacillus acidophilus, as potential probiotics, on the immune response and resistance of Tilapia nilotica (Oreochromis niloticus) to challenge infections. Fish Shellfish Immunol., 2008b, 25: 128-136
Aly S M, Mohamed M F, John G. Effect of probiotics on the survival, growth and challenge infection in Tilapia nilotica (Oreochromis niloticus). Aquaculture Research 2008c, 39: 647-656
Amann R I, Ludwig W, Schleifer K H. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev.,1995, 59: 143-169
Anderson D P, Siwicki A K, Rumsey G L. Injection or immersion delivery of selected immunostimulants to trout demonstrate enhancement of nonspecific defense mechanisms and protective immunity. Diseases in Asian Aquaculture, Vol.Ⅱ. Fish Health Section, Manila: Asia Fisheries Society, 1995, 413-426
Anderson D P, Siwicki A K. Duration of protection against Aeromonas salmonicida in brook trout immunostimulated with glucan or chitosan by injection or immersion. Pro. Fish Cult., 1994, 56: 258-261
Anderson D P. Immunostimulants, adjuvants, and vaccine carriers in fish: application to aquaculture. Fish Diseases, 1992(2): 281-307
Apún-Molina J P, Santamaría-Miranda A, Luna-Gonzalez A, et al. Effect of potential probiotic bacteria on growth and survival of tilapia Oreochromis niloticus L., cultured in the laboratory under high density and suboptimum temperature. Aquaculture Research, 2009, 40: 887- 894
Bagheri T, Hedayati S A, Yavari V, et al. Growth, Survival and Gut Microbial Load of Rainbow Trout (Onchorhynchus mykiss) Fry Given Diet Supplemented with Probiotic during the Two Months of First Feeding. Turkish Journal of Fisheries and Aquatic Sciences, 2008, 8: 43-48
Bagni M, Romano N, Finoia M G,et al. Short- and long-term effects of a dietary yeastβ-glucan (Macrogard) and alginic acid (Ergosan) preparation on immune response in sea bass (Dicentrarchus labrax). Fish Shellfish Immunol., 2005, 18: 311-325
Balcázar J L, de Blas I, Ruiz-Zarzuela I, et al. The role of probiotics in aquaculture. Veterinary Microbiology, 2006, 114: 173-186
Balcázar J L, Rojas-Luna T, Cunningham D P. E147-150
Balcázar J L. Evaluation of probiotic bacterial strains in Litopenaeus vannamei. Final Report, National Center for Marine and Aquaculture Research, Guayaquil, Ecuador. 2003
Bandyopadhyay P, Mohapatra P K D. Effect of a probiotic bacterium Bacillus circulans PB7 in the formulated diets: on growth, nutritional quality and immunity of Catla catla (Ham.) Fish Physiol. Biochem., 2009, 35: 467-478
Barracco M A, Duvic B, Soderhall K. Theβ-1,3-glucan binding protein from the crayfish Pacifastacus leniusculus, when reacted with aβ-1,3-glucan induces spreading and degranulation of crayfish granular cell. Cell Tissue Res, 1991, 266: 491-497
Bondad-Reantaso M, Subasinghe R P, Arthur J R, et al. Disease and health management in Asian aquaculture. Vet. Parasitology, 2005, 132: 249-272
Boolootian R A., Giese A C. Clotting of echinoderm coelomic fluid. J. Exper. Zool., 1959, 140:207-211
Boonyaratpalin S, Boonyyaratpalin M, Supamattaya K, et al. Effects of peptidoglycan (PG) on growth, survival, immune response and tolerance to stress in black tiger shrimp, Penaeus monodon. In: Diseases in Asian Aquaculture. Vol. 11. Fish Health Section, Asian Fisheries Society, Manila: Philippines, 1995, 469-477
Bornet F R, Brouns F. Immune-stimulating and gut health-promoting properties of short chain fructo-oligosaccharides. Nutr. Rev., 2002, 60(11): 326-334
Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Analytical Biochemistry, 1976, 72: 248-254
Brown G D, Gordon S. Fungalβ-Glucans and Mammalian Immunity. Immunity, 2003, 19, 311-315.
Buentello J A, Neill W H, Gatlin D M III, Effects of dietary prebiotics on the growth, feed efficiency and non-specific immunity of juvenile red drum Sciaenops ocellatus fed soybean-based diets. Aquaculture Research, 2010,41: 411-418
Bulgakov A A, Eliseikina M G, Petrova I Y, et al. Molecular and biological characterization of a mannan-binding lectin from the holothurian Apostichopus japonicus. Glycobiology,2007, 17: 1284-1288
Byun J W, Park S C, Benno Y, et al. Probiotic effect of Lactobacillus sp. DS-12 in ?ounder (Paralichthys olivaceus). J. Gen. Appl. Microbiol., 1997, 43: 305–308
Campa-Córdova A I, Hernández-Saavedra N Y, De Philippis R, et al. Generation of superoxide anion and SOD activity in haemocytes and muscle of American white shrimp(Litopenaeus vannamei) as a response toβ-glucan and sulphated polysaccharide. Fish Shellfish Immunol., 2002, 12: 353-366
Canicatti C, D'Ancona G, Cellular aspects of Holothuria polii immune response. J. Invertebr. Pathol., 1989, 53: 152-158
Canicatti C, G(o|¨)tz P. Dopa oxidation by Holothuria polii coelomocyte lysate. J. Invertebrate Pathol., 1991, 58: 305-310
Canicatti C, Lysosomal enzyme pattern in Holothuria polii coelomocytes. J. Invert Pathol., 1990, 56: 70-74
Canicatti C, Pagliara P, Stabili L. Sea urchin coelomic fluid agglutinin mediates coelomocyte adhesion. Eur. J. Cell Biol., 1992, 58(2): 291-295
Canicatti C, Parrinello N. Hemaglutinin and hemolysin level in coelomic fluid from Holothuria polii (Echinodermata) following sheep erythrocyte injection. Biol. Bull., 1985, 168: 175-182
Canicatti C, Rizzo A. A 220 kDa coelomocyte aggregating factor involved in Holothuria polii celluar clotting. Eur. J. cell Biol., 1991, 56: 79-83
Canicatti C, Seymour J. Evidence for phenoloxidase activity in Holothuria tubulosa (Echinodermata) brown bodies and cells. Parasitol. Res., 1991, 77: 50-53
Canicatti C. Hemolysins: pore-forming proteins in invertebrates. Celluar and Molecular life science, 1990, 46(3): 239-244
Cao Q Z, Lin Z B. Antitumor and anti-angiogenic activity of Ganoderma lucidum polysaccharides peptide. Acta Pharmacol. Sin., 2004, 25: 833-838
Castex M, Lemaire P, Wabete N, et al. Effect of probiotic Pediococcus acidilactici on antioxidant defences and oxidative stress of Litopenaeus stylirostris under Vibrio nigripulchritudo challenge. Fish Shellfish Immunol., 2010, 28: 622-631
Chang C F, Chen H Y, Su M S,et al. Immunomodulation by dietaryβ-1,3-glucan in the brooders of the black tiger shrimp Penaeus monodon Fish Shellfish Immunolo., 2000, 10: 505-514
Chang C F, Su M S, Chen H Y, et al. Dietaryβ-1,3-glucan effectively improves immunity and survival of Penaeus monodon challenged with white spot syndrome virus. Fish Shellfish Immunol., 2003, 15: 297-310
Chang Y Q, Yu C Q, Song X. Pond culture of sea cucumbers, Apostichopus japonicus, in Dalian. In: Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy, 2004. 269-272
Chen J X. Present status and prospects of sea cucumber industry in China. In: Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy, 2004. 25-38
Chia F S, Xing J. Echinoderm coelomocytes, a review. Zool. Stud., 1996, 35(4): 231-254 Chythanya R, Karunasager I, Karunasager I. Inhibition of shrimp pathogenic vibirios by a marine PseudomonasⅠ-2 strain. Aquaculture, 2002, 208: 1-10
Coffaro K A, Hinegardner A T. Immune response in the sea urchin Lytechinus oictus Science, 1977, 197: 1389-1390
Cook M T, Hayball P J, Hutchinson W, et al. Administration of a commercial immunostimulant preparation, EcoActiva as a feed supplement enhances macrophase respiratory burst and the growth rate of snapper (Pagrus auratus, Sparidae (Bloch and Schneider)) in winter. FishShellfish Immunol., 2003, 14: 333-345
Coteur G, Warnau M, Jangoux M, Dubois P. Reactive oxygen species (ROS) production by amoebocytes of Asterias rubens (Echinodermata). Fish Shellfish Immunol., 2002, 12(3): 187-200
Couso N, Castro R, Magarinos B, et al. Effect of oral administration of glucans on the resistance of gilthead seabream to pasteurellosis. Aquaculture, 2003, 219: 99-109
Culjak V, Bogut G, Has-Schon E, Milakovic Z, Canecki K (2006) Effect of Bio-Mos on performance and health of juvenile carp. In: Nutrition and biotechnology in the feed and food industries: Alltech’s 22nd annual symposium (suppl. 1-abstracts of posters presented), Lexington, KY, USA, 2006
Dalmin G, Kathiresan K, Purushothaman A. Effect of probiotics on bacterial population and health status of shrimp in culture pond ecosystem. Indian J. Exp. Biol., 2001, 39: 939-942
Dalmo R A, B(?)gwald J.β-glucans as conductors of immune symphonies. Fish Shellfish Immunol., 2008,25: 384-396
Daniels C L, Merrifield D L, Boothroyd D P, et al. Effect of dietary Bacillus spp. and mannan oligosaccharides (MOS) on European lobster (Homarus gammarus L.) larvae growth performance, gut morphology and gut microbiota. Aquaculture, 2010, 304: 49-57
Deeseenthum S, Leelavatcharamas V, Brooks J D. Effect of feeding Bacillus sp. As probiotic bacteria on growth of giant freshwater prawn (Macrobrachium rosenbergii de Man). Pakistan Journal of Biological Sciences, 2007, 10(9): 1481-1485
Delzenne N, Aertssens J, Verplaetse H, et al. Effect of fermentable fructo-oligosaccharides on mineral, nitrogen and energy digestive balance in the rat. Life Sci., 1995, 57: 1579-1587
Didier J, Philippe D., Michel J. Defensive mechanisms of holothriod(Echinodermata): Formation, role and fate of intracoelomic brown bodies in sea cucumber Holothuria tubulosa. Cell and Tissue Research, 1996, 283(1): 99-106
Dolmatova L S, Eliseikina M G, Romashina V V. Antioxidant enzymatic activity of coelomocytes of the far East sea cucumber Eupentacta fraudatrix. J. Evol. Biochem. Physiol., 2004, 40(2): 126-135
Du X J, Zhao X F, Wang J X. Molecular cloning and characterization of a lipopolysaccharide and β-1,3-glucan binding protein from fleshy prawn (Fenneropenaeus chinensis). Molecular Immunology 2007, 44: 1085-1094
Dybas L, Fankboner P V. Holothurian survival strategies: mechanisms for the maintenance of a bacteriostatic environment in the coelomic cavity of the sea cucumber, Parastichopus californicus. Dev. Comp. Immunol., 1986, 10(3): 311-330
El-Dakar A Y, Shalaby S M, Saoud I P. Assessing the use of a dietary probiotic/prebiotic as an enhancer of spinefoot rabbitfish Siganus rivulatus survival and growth. Aquaculture Nutrition, 2007, 13: 407-412
Eliseikina M G, Magarlamov T Y. Coelomocyte morphology in the holothurians Apostichopusjaponicus and Cucumaria japonica. Russian Journal of Marine Biology, 2002, 28(3): 197-202
El-Rhman A M A, Khattab Y A E, Shalaby A M E. Micrococcus luteus and Pseudomonas species as probiotics for promoting the growth performance and health of Nile tilapia, Oreochromis niloticus. Fish Shellfish Immunol., 2009, 27: 175-180
Engstad R E, Robertsen B. Specificity of aβ-glucan receptor on macrophages from Atlantic salmon (Salmo salar L.). Dev. Comp. Immunol., 1994, 18: 397-408
FAO/WHO. Joint FAO/WHO (Food and Agriculture Organization/World Health Organization) working group report on drafting guidelines for the evaluation of probiotics in food. London,
Ontario, Canada. guidelines for the evaluation of probiotics in food. Joint working group report on drafting. London, Ontario, 2002, pp. 1-11.
Farzanfar A. The use of probiotics in shrimp aquaculture. FEMS Immunol Med Microbiol. 2006, 48: 149-158
Fontaine A R., Lambert P. The fine structure of the leucocytes of the holothurian, Cucumaria miniata. Can. J. Zool., 1977, 55: 1530-1544
Franchini A, Conte A, Ottaviani E. Nitric oxide: an ancestral immunocyte effector molecule. Advances in Neuroimmunology, 1995, 5: 463-478
Fu L L, Shuai J B, Xu Z R, et al. Immune responses of Fenneropenaeus chinensis against white spot syndrome virus after oral delivery of VP28 using Bacillus subtilis as vehicles. Fish Shellfish Immunol., 2010, 28: 49-55
Fuller R. Probiotic in man and animals. J. Appl. Bacteriol., 1989, 66: 365-378
Gatesoupe F J. The use of probiotics in aquaculture, Aquaculture, 1999, 180: 147-165
Ghosh K, Sen S K, Ray A K. Supplementation of an isolated fish gut bacterium, Bacillus circulans, in formulated diets for rohu, Labeo rohita, fingerlings. Israeli J. Aquacult. Bamidgeh, 2003, 55: 13-21
Gibson G R, Roberfroid M B. Dietary modulation of the human colonic microbiota: introducing the concept of prebiotics. J. Nutr., 1995, 125: 1401-1412
Gildberg A, Mikkelsen H, Sandaker E, et al. Probiotic effect of lactic acid bacteria in the feed on growth and survival of fry of Atlantic cod (Gadus morhua). Hydrobiologia 1997, 352: 279-285
Goldin B R. Health benefits of probiotics. Br. J. Nutr., 1998, 80: S203-207
Gomez-Gil B, Roque A, Turnbull J F. The use and selection of probiotic bacteria for use in the culture of larval aquatic organisms. Aquaculture, 2000, 191: 259-270
Gopalakannan A, Arul V. Immunomodulatory effects of dietary intake of chitin, chitosan and levamisole on the immune system of Cyprinus carpio and control of Aeromonas hydrophila infection in ponds. Aquaculture, 2006, 255: 179-187
Gowda N M, Goswami U, Khan M I. Purification and characterization of a T-antigen specific lectin from the coelomic fluid of a marine invertebrate, sea cucumber (Holothuria scabra).Fish Shellfish Immunol., 2008, 24(4):450-458
Gram L, Melchiorsen J, Spanggaard B, et al. Inhibition of Vibrio anguillarum by Pseudomonasfluorescens strain AH2, a possible probiotic treatment of fish. Appl. Environ. Microbiol.,1999, 65, 969-973
Grisdale-Helland B, Helland S J, Gatlin D M III. The e ects of dietary supplementation withmannanoligosaccharide, fructooligosaccharide or galsctooligosaccharide on the growth andfeed utilization of Atlantic salmon (Salmo salar L.). Aquaculture, 2008, 283: 163-167
Gu M, Ma H M, Mai K S, Zhang W B, Ai Q H, Wang X J, Bai N. Immune response of seacucumber Apostichopus japonicus coelomocytes to several immunostimulants in vitro.Aquaculture, 2010, 306: 49-56
Gullian M, Thompson F, Rodriguez J. Selection of probiotic bacteria and study of theirimmunostimulatory effect in Penaeus vannamei. Aquaculture, 2004, 233: 1-14
Hai N V, Buller N, Fotedar R. Effects of probiotics (Pseudomonas synxantha and Pseudomonasaeruginosa) on the growth, survival and immune parameters of juvenile western king prawns(Penaeus latisulcatus Kishinouye, 1896). Aquaculture Research, 2009, 40: 590-602
Hatakeyama T, Nagatomo H, Yamasaki N. Interaction of the hemolytic lectin CEL-III from themarine invertebrate Cucumaria echinata with the erythrocyte membrane. J. Biol. Chem.,1995, 270(8): 3560-3564.
Hatakeyama T, Sato T, Taira E, et al. Characterization of the interaction of hemocytic lectin CEL-Ⅲfrom the marine invertebrate, cucumaria echinata,with artificial lipid membranes:involvement of neutral sphingoglycolipids in the pore-forming process. J Biochem(Tokyo),1999, 125(2): 277-284
Haug T, Kjuul A K, Styrvold O B, et al. Antibacterial activity in Strongylocentrotus droebachiensis(Echinoidea), Cucumaria frondosa (Holothuroidea), and Asterias rubens (Asteroidea). J.Invertebr. Pathol., 2002, 81(2):94-102
Hernández-López J, Gollas-Galvan T, Vargas-Albores F. Activation of the prophenoloxidasesystem of brown shrimp (Penaues californiensis Holmes) Comp. Biochem. Physiol., 1996,Part C, 113: 61-66
Hjelm M, Bergh ?, Riaza A, et al. Selection and identification of autochthonous potentialprobiotic bacteria from turbot larvae (Scophthalmus maximus) rearing units. Syst. Appl.Microbiol., 2004, 27: 360-371
Hua X M, Zhou H Q, Zhang D Q, et al. Effect of dietary chitosan and probiotics on diseaseresistance and immunity of obscure puffer (Fugu obscures) Journal of fisheries of China,2007, 31(4): 478-486
Huang C C, Song Y L. Maternal transmission of immunity to white spot syndrome associatedvirus (WSSV) in shrimp (Panaeus monodon). Dev. Comp. Immunol, 1999, 23: 545-552
Ibrahema M D, Fathi M, Mesalhy S, et al. Effect of dietary supplementation of inulin and vitaminC on the growth, hematology, innate immunity, and resistance of Nile tilapia (Oreochromisniloticus). Fish Shellfish Immunol., 2010, 29: 241-246
Isaeva V V, Korenbaun E S. Defense functions of coelomocytes and immunity of echinoderms. Soviet Journal of Marine Biology, 1990, (15): 353-363
Itami T, Asano M, Tokushige K, et al. Enhancement of disease resistance of kuruma shrimp, Penaeus japonicus, after oral administration of peptidoglycan derived from Bifidobacterium thermophilum. Aquaculture, 1998, 164: 277-288
Itami T, Kondo M, Uozu M, et al. Enhancement of resistance against Enterococcus seriolicida infection in yellowtail, Seriola quinqueradiata (Temninck and Schlegel), by oral administration of peptidoglycan derived from Bifidobacterium thermophilum. J Fish Dis, 1996, 19: 185-187
Jayaraj S S, Thiagarajan R, Arumugam M, et al. Isolation, purification and characterization of β-1,3-glucan binding protein from the plasma of marine mussel Perna viridis. Fish Shellfish Immunol., 2008, 24, 715-725
Jonathan P R, Smith L C, Mariano L C, et al. Genomic insights into the immune system of the sea urchin. Science, 2006, 314: 952-956
Karp R D, Hiloernann W H. Specific allograft reactivity in the sea star Dermasterias imbricate. Transplantation, 1976, 22: 434-439
Kesarcodi-Watson A, Kaspar H, Lategan M J, et al. Probiotics in aquaculture: The need, principles and mechanisms of action and screening processes. Aquaculture, 2008, 274: 1-14
Kodama H, Hirota Y, Mukamoto M, et al. Activation of rainbow trout (Oncorhynchus mykiss) phagocytes by muramyl dipeptide. Dev. Comp. Immunol., 1993, 7: 129-140
Kono T, Ponpornpisit A, Sakai M. The analysis of expressed genes in head kidney of common carp Cyprinus carpio L. stimulated with peptidoglycan. Aquaculture, 2004, 235: 37-52
Kono T, Sakai M. The analysis of expressed genes in the kidney of Japanese flounder, Paralichthys olivaceus, injected with the immunostimulant peptidoglycan. Fish Shellfish Immunol., 2001, 11: 357-366
Kozasa M. Toyocerin (Bacillus toyoi) as growth promotor for animal feeding. Microbiologie Aliments Nutrition, 1986, 4(1): 121-135
Kumar B P, Sahu N P, Saharan N, et al. Effect of dietary source and level of chitin on growth and survival of post-larvae Macrobrachium rosenbergii. J. Appl. Ichthyol. 2006, 22: 363-368
Kumar R, Mukherjee S C, Ranjan R, et al. Enhanced innate immune parameters in Labeo rohita (Ham.) following oral administration of Bacillus subtilis. Fish Shellfish Immunol., 2008, 24: 168-172
Kwak J K, Park S W, Koo J G, et al. Enhancement of the non-specific defense activities in carp (Cyprinus carpio) and flounder (Paralichthys olivaces) by oral administration of Schizophyllan. Acta Biotechnol, 2003, 23(4): 359-371
Li J Q, Tan B P, Mai K S. Dietary probiotic Bacillus OJ and isomaltooligosaccharides in?uence the intestine microbial populations, immune responses and resistance to white spot syndromevirus in shrimp (Litopenaeus vannamei). Aquaculture, 2009, 291: 35-40
Li J Y, Sun X Q, Zheng F R, et al. Screen and effect analysis of immunostimulants for sea cucumber, Apostichopus japonicas. Chinese Journal of Oceanology and Limnology, 2009, 27(1): 80-84
Li P, Burr G S, Gatlin D M III, et al. Dietary Supplementation of Short-Chain Fructooligosaccharides In?uences Gastrointestinal Microbiota Composition and Immunity Characteristics of Pacific White Shrimp, Litopenaeus vannamei, Cultured in a Recirculating System. The Journal of Nutrition, 2007, 137: 2763-2768
Li P, Wen Q, Gatlin D M III. Dose-dependent in?uences of dietaryβ-1,3-glucan on innate immunity and disease resistance of hybrid striped bass Morone chrysops×Morone saxatilis. Aquaculture Research, 2009, 40: 1578-1584
Lilley D M, Stillwell R J. Probiotics: growth promoting factors produced by micro-organisms. Science, 1965, 147: 747-748
Lin Y C, Vaseeharan B, Chen J C. Identification and phylogenetic analysis on lipopolysaccharide andβ-1,3-glucan binding protein (LGBP) of kuruma shrimp Marsupenaeus japonicus. Developmental and Comparative Immunology 2008, 32: 1260-1269
Liu F S, Li F H, Dong B, et al. Molecular cloning and characterisation of a pattern recognition protein, lipopolysaccharide andβ-1,3-glucan binding protein (LGBP) from Chinese shrimp Fenneropenaeus chinensis. Mol. Biol. Rep. 2009, 36: 471-477
López N, Cuzon G, Gaxiola G, et al. Physiological, nutritional, and immunological role of dietary β-1,3-glucan and ascorbic acid 2-monophosphate in Litopenaeus vannamei juveniles. Aquaculture, 2003, 224, 223-243
Lv H Y, Zhou Z G, Rudeaux F, et al. Effects of dietary short chain fructooligosaccharides on intestinal microflora, mortality and growth performance of Oreochromis aureus×O. niloticus. Chin. Anim. Sci., 2007, 19(6): 691-697
Mahious A S, Gatesoupe F J, Hervi M, et al. Effects of dietary inulin and oligosaccharides as prebiotics for weaning turbot Psetta maxima (Linneuas, C. 1758). Aquacult. Int., 2006, 14: 219-229
Manning T S, Gibson G R. Prebiotics. Best Pract. Res. Clin Gastroenterol, 2004, 18: 287-298
Martin F P, Collen M C. Bias in template-to-product ratios in multitemplate PCR. Appl. Environ. Microbiol., 1998, 64: 3724-3730
Matsui T, Ozeli Y, Suzuli M, et al. Purification and characterization of two Ca2+ dependent lectins from coelomic plasma of sea cucumber, Stichopus japonicus. J. Biochem., 1994, 116(5): 1127-1133
Meng Z, Shao J, Xiang L. CPG oligodeoxynucleotides activate grass carp (Ctenopharyngodon idellus) macrophages. Dev. Comp. Immunol. 2003, 27(4): 313-321
Merrifield D L, Dimitroglou A, Bradley G., et al. Probiotic applications for rainbow trout (Oncorhynchus mykiss Walbaum) I. Effects on growth performance, feed utilization,intestinal microbiota and related health criteria. Aqua. Nutr., 2010, 16: 504-510
Misra C K, Das B K, Mukherjee S C, et al. Effect of long term administration of dietaryβ-glucan on immunity, growth and survival of Labeo rohita fingerlings. Aquaculture, 2006b, 255: 82-94
Misra C K, Das B K, Mukherjee S C, et al. Effect of multiple injections ofβ-glucan on non-specific immune response and disease resistance in Labeo rohita fingerlings. Fish Shellfish Immunol., 2006a, 20: 305-319
Nayak S K, Swain P, Mukherjee S C. Effect of dietary supplementation of probiotic and vitamin C on the immune response of Indian major carp, Labeo rohita (Ham.) Fish Shellfish Immunol. 2007, 23: 892-896
Nayak S K. Probiotics and immunity: a fish perspective. Fish Shellfish Immunol., 2010, 29: 2-14
Newaj-Fyzul A, Adesiyun A A, Mutani A. et al. Bacillus subtilis AB1 controls Aeromonas infection in rainbow trout (Oncorhynchus mykiss, Walbaum). Journal of Applied Microbiology, 2007, 103: 1699-1706
Olivier G, Evelyn T, Lallier R. Immunity to Aeromoas salmonicida in coho salmon (Oncorhynchus kisutch) induced by modified Freund’s complete adjuvant: its non-specific nature and the probable role of mocrophages in the phenomenon. Dev. Comp. Immunol., 1985, 9: 419-432
Olsson C, Westerdahl A, Conway P L, et al. Intestinal colonization potential of turbot (Scophthalmus maximus L.) and dab(Limanda limanda)associated bacteria with inhibition effects against vibrio anguillarum. Appl. Environ. Micro., 1992, 58(3): 551-556
Pan, X, Wu, T, Song Z, et al. Immune responses and enhanced disease resistance in Chinese drum, Miichthys miiuy (Basilewsky), after oral administration of live or dead cells of Clostridium butyrium CB2. Journal of Fish Diseases, 2008, 31: 679-686
Parker R B. Probiotics, the other half of the antibiotics story. Anim. Nutr. Health, 1974, 29: 4-8
Pedro G S, César A B, Francisco R, et al. Lipopolysaccharides induce intestinal serum amyloid A expression in the sea cucumber Holothuria glaberrima. Dev. Comp. Immunol., 2003, 27(2): 105-110.
Powell A, Rowley A F. The effect of dietary chitin supplementation on the survival and immune reactivity of the shore crab, Carcinus maenas. Comparative Biochemistry and Physiology, 2007, 147(A): 122-128
Qi Z Z, Zhang X H, Boon N, et al. Probiotics in aquaculture of China -current state, problems and prospect. Aquaculture, 2009, 290: 15-21
Ratcliffe N A, Rowley A F. Ivertebrate blood cells. New York:Academic press New York, 1980. 513-526
Rattanachai A, Hirono I, Ohira,T, et al. Peptidoglycan inducible expression of a serine proteinase homologue from kuruma shrimp (Marsupenaeus japonicus). Fish Shellfish Immunol., 2005, 18: 39-48
Rengpipat S, Phianphak W, Piyatiratitivorakul S, et al. Effects of a probiotic bacterium on black tiger shrimp Penaeus monodon survival and growth. Aquaculture, 1998, 167: 301-313
Rengpipat S, Rukpratanporn S, Piyatiratitivorakul S, et al. Immunity enhancement in black tiger shrimp (Penaeus monodon) by a probiont bacterium (Bacillus S11). Aquaculture, 2000, 191: 271-288
Ring(?) E, Olsen T (?), Gifstad R A, et al. Prebiotic in aquaculture: a review. Aquaculture Nutrition, 2010,16: 117-136
Roch P, Canicatti ., Sammarco S. Tetrameric structure of the active phenoloxidase evidence in the coelomocytes of the echinoderm Holothuria tubulosa. Comp. Biochem. Physiol., 1992, 102B: 349-355
Rodríguez-Ramos T, Carpio Y, Bolívar J, et al. An inducible nitric oxide synthase (NOS) is expressed in hemocytes of the spiny lobster Panulirus argus: Cloning, characterization and expression analysis. Fish Shellfish Immunol., 2010, 29: 469-479
Rodríguez-Ramos T, Espinosa E, Hernández-López J, et al. Effects of Echerichia coli lipopolysaccharides and dissolved ammonia on immune response in southern white shrimp Litopenaeus schmitti. Aquaculture, 2008, 274: 118-125
Sahoo P K, Das A, Mohanty S. et al. Dietaryβ-1, 3-glucan improves the immunity and disease resistance of freshwater prawn Macrobrachium rosenbergii challenged with Aeromonas hydrophila. Aquaculture. Research, 2008, 39, 1574-1578
Sahoo P K, Mukherjee S C. The effect of dietary immunomodulation upon E.tarda vaccination in healthy and immunocompromised Indian Major Carp (L.rohita). Fish Shellfish Immunol., 2002, 12(1): 1-16
Sakai M. Current research statue of fish immunostimulant. Aquaculture, 1999, 172: 63-92
Salinas I, Cuesta A, Esteban M A, et al. Dietary administration of Lactobacillus delbrueckii and Bacillus subtilis, single or combined, on gilthead seabream cellular innate immune responses. Fish Shellfish Immunol., 2005 19: 67-77
Sealey W M, Barrows F T, Hang A, et al. Evaluation of the ability of barley genotypes containing different amounts ofβ-glucan to alter growth and disease resistance of rainbow trout Oncorhynchus mykiss. Animal Feed Science and Technology, 2008, 141: 115-128
Selvaraj V, Sampath K, Sekar V. Administration of yeast glucan enhances survival and some non-specific and specific immune parameters in carp (Cyprinus carpio) infected with Aeromonas hydrophila Fish Shellfish Immunol., 2005, 19(4): 293-306
Sghir A, Chow J M, Mackie R I. Continuous culture selection of bifidobacteria and lactobacilli from human fecal samples using fructooligosaccharide as selective substrate. J. Appl. Microbiol., 1998, 85: 769-777
Sharifuzzaman S M, Austin B. In?uence of probiotic feeding duration on disease resistance and immune parameters in rainbow trout. Fish Shellfish Immunol., 2009, 27: 440-445
Sisak F. Stimulation of phagocytosis as assessed by luminol-enhanced chemiluminescence andresponse to salmonella challenge of poultry fed diets containing mannan oligosaccharides. In: Lyons T.P., Jacqus K.A.(Eds.). Biotechnology in the feed industry, proceedings of Alltech’s 10th annual symposium. Nottingham, UK: Nottingham University Press. 1995
Siwicki A K, Anderson D P , Rumsey GL. Dietary intake of immunostimulants by rainbow trout affects non-specific immunity and protection against furunculosis.Vet Immunopathol, 1994, 41: 125-139
Smith L C, Britten R J, Davidson E H. Sea urchin genes expressed in activated coelomocytes are identified by expressed sequence tags. Complement homologues and other putative immune response genes suggest immune system homology within the deuterostomes. Journal of Immunology. 1996, 156: 593-602
Smith L C, Britten R J, Davidson E H. SpCoel1: A sea urchin profilin gene expressed specifically in coelomocytes in response to injury. Molecular Biology of the Cell, 1992, 3: 403-414
Smith L C, Clow L A, Terwilliger D P. The ancestral complement system in sea urchins. Immunol. Rev. 2001, 180:16-34
Smith P, Davey S. Evidence for the competitive excluding of Aeromonas salmonicida from fish with stress inducible furunculosis by a fluorescent pseudomonad. J. of Fish Disease, 1993, 16: 521-524
Smith V J. The echinoderms. In: Rafcliffe N A, Rowley A F (eds). Invertebrate blood cells, London: Academic Press, 1981. 513-562
Sogaard H. Microbials for feed: Beyond lactic acid bacteria. Feed international, 1990, 11: 22-37
Son V M, Chang C C, Wu M C, et al. Dietary administration of the probiotic, Lactobacillus plantarum, enhanced the growth, innate immune responses, and disease resistance of the grouper Epinephelus coioides. Fish Shellfish Immunol., 2009, 26: 691-698
Song H L, Hsieh Y T. Immunostimulation of tiger shrimp (Penaeus monodon) hemocytes for generation of microbicidal substances: analysis of reactive oxygen species. Dev. Comp. Immunol, 1994, 18(3): 201-209
Song Y L, Yu C I, Lien T W, et al. Haemolymph parameters of Pacific white shrimp (Litopenaeus vannamei) infected with Taura syndrome virus. Fish Shellfish Immunol., 2003, 14: 317-331
Sritunyalucksana K, Lee S Y, Soderhall K A.β-1,3-glucan binding protein from the black tiger shrimp Penaeus monodon. Dev. Comp. Immunol., 2002, 26: 237-245
Sritunyalucksana K, Sithisarn P, Withayachumnarnkul B, et al. Activation of prophenoloxidase, agglutinin and antibacterial activity in haemolymph of the black tiger prawn, Penaeus monodon, by immunostimulants. Fish Shellfish Immunol., 1999, 9: 21-30
Staykov Y, Denev S, Spring P. In?uence of dietary mannan oligosaccharides (Bio-Mos) on growth rate and immune function of common carp (Cyprinus carpio L.). In: Howell B, Flos R (eds) Lessons from the past to optimise the future. European Aquaculture Society, Special Publication No 35, 2005, pp 431-432
Staykov Y, Spring P, Denev S, et al. Effect of a mannan oligosaccharide on the growth performanceand immune status of rainbow trout (Oncorhynchus mykiss). Aquacult. Int., 2007, 15: 153-161
Sugita H, Hirose Y, Matsuo N, et al. Production of the antibacterial substance by Bacillus sp.strain NM 12, an intestinal bacterium of Japanese coastal fish. Aquaculture, 1998, 165(3-4): 269-280
Sun H L, Liang M Q, Yan J P, et al. Nutrient requirements and growth of the sea cucumber, Apostichopus japonicus In: Lovatelli A. (Eds.), Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy. 2004. pp. 327-331
Sun Y Z, Yang H L, Ma R L, et al. Probiotic applications of two dominant gut Bacillus strains with antagonistic activity improved the growth performance and immune responses of grouper Epinephelus coioides. Fish Shellfish Immunol., 2010, 29: 803-809
Sun Y, Jin L, Wang T. Polysaccharides from Astragalus membranaceus promote phagocytosis and superoxide anion (O2-) production by coelomocytes from sea cucumber Apostichopus japonicus in vitro. Comp. Biochem. Physiol., 2008, 147(C):2 93-298
Suzer C, (?)oban D, , Kamaci H O, et al. Lactobacillus spp. bacteria as probiotics in gilthead sea bream (Sparus aurata, L.) larvae: Effects on growth performance and digestive enzyme activities. Aquaculture, 2008, 280: 140-145
Swain S M, Singh C, Arul V. Inhibitory activity of probiotics Streptococcus phocae PI80 and Enterococcus faecium MC13 against Vibriosis in shrimp Penaeus monodon. World J. Microbiol. Biotechnol., 2009, 25: 697-703
Sweetman J W, Torrecillas S, Dimitroglou A, et al. REVIEW ARTICLE Enhancing the natural defences and barrier protection of aquaculture species. Aquaculture Research, 2010, 41, 345-355
Taoka Y, Maeda H, Jo J Y, et al. Growth, stress tolerance and non-specific immune response of Japanese flounder Paralichthys olivaceus to probiotics in a closed recirculating system. Fish Sci., 2006, 72, 310-321
Tlaskalová-HogenováH, StepánkováR, Hudcovic T, et al. Commensal bacteria (normal microflora), mucosal immunity and chronic inflammatory and autoimmune diseases. Immunol. Lett., 2004, 93: 97-108
Torrecillas S, Makol A, Caballero M J, et al. Immune stimulation and improved infection resistance in European sea bass (Dicentrarchus labrax) fed mannan oligosaccharides. Fish Shellfish Immunol., 2007, 23: 969-981
Tseng D Y, Ho P L, Huang S Y, et al. Enhancement of immunity and disease resistance in the white shrimp, Litopenaeus vannamei, by the probiotic, Bacillus subtilis E20. Fish Shellfish Immunol., 2009, 26: 339-344
Vargas-Albores F, Yepiz-Plascencia G. Beta glucan binding protein and its role in shrimp immune response. Aquaculture, 2000, 191, 13-21
Vaseeharan B, Ramasamy P. Control of pathogenic Vibrio spp. by Bacillus subtilis BT23, apossible probiotic treatment for black tiger shrimp Penaeus monodon. Letters in applied microbiology, 2003, 36: 83-87
Vendrell D, Balcázar J L, de Blas, I. et al. Protection of rainbow trout (Oncorhynchus mykiss) from lactococcosis by probiotic bacteria. Comparative Immunology, Microbiology and Infectious Diseases, 2008, 31: 337-345
Venkat H K, Sahu N P, Jain K K. Effect of feeding Lactobacillus-based probiotics on the gut micro?ora, growth and survival of postlarvae of Macrobrachium rosenbergii (de Man). Aquaculture Research, 2004, 35: 501-507
Verschuere L, Rombaut G, Sorgeloos P, et al. Probiotic bacteria as biological control agents in aquaculture. Microbiology and Molecular Biology Review, 2000, 64: 655-671
Wang S H, Chen J C. The protective effect of chitin and chitosan against Vibrio alginolyticus in white shrimp Litopenaeus vannamei Fish Shellfish Immunol., 2005, 19: 191-204
Wang Y B, Han J Z. The role of probiotic cell wall hydrophobicity in bioremediation of aquaculture. Aquaculture, 2007, 269: 349-354
Wang Y B, Li J R, Lin J. Probiotics in aquaculture: Challenges and outlook Aquaculture, 2008, 281: 1-4
Wang Y B. Effect of probiotics on growth performance and digestive enzyme activity of the shrimp Penaeus vannamei. Aquaculture, 2007, 269: 259-264
Wang Y C, Chang P S, Chen H Y. Differential time-series expression of immune-related genes of Pacific white shrimp Litopenaeus vannamei in response to dietary inclusion of β-1,3-glucan.Fish Shellfish Immunol., 2008, 24: 113-121
Wang Y G, Zhang C Y, Rong X J, et al. Diseases of cultured sea cucumber, Apostichopus japonicus, in China. In: Lovatelli A. (Ed.), Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy. 2004, pp. 297-310
Wang Y J, Osatomi K, Yoshida A, et al. Extracellular products from virulent strain of Edwardsiella tarda stimulate mouse macrophages (RAW264.7) to produce nitric oxide (NO) and tumor necrosis factor (TNF)-α. Fish Shellfish Immunol., 2010, 29: 778-785
Whittington R, Lim C, Klesius P H. Effect of dietaryβ-glucan levels on the growth response and efficacy of Streptococcus iniae vaccine in Nile tilapia, Oreochromis niloticus. Aquaculture, 2005, 248: 217-225
Xing J, chia F S. Opsonin-like molecule found in coelomic fluid of sea cucumber, Holothuria leucospilota. Marine Biology, 2000, 136: 979-986
Xing J, Leung M F, Chia F S. Quantitative analysis of phagcytosis by amoebocytes in a sea cucumber Holothuria leucospilota. Invert. Biol., 1998, 117: 67-74
Yeh F C, Wu S H, Lai C Y, et al. Demonstration of nitric oxide synthase activity in crustacean hemocytes and anti-microbial activity of hemocyte-derived nitric oxide. Comparative Biochemistry and Physiology, 2006, Part B 144(1):11-17
Yousefian M, Amiri M S, A review of the use of prebiotic in aquaculture for fish and shrimp. African Journal of Biotechnology, 2009, 8(25): 7313-7318
Zhang L, Mai K S, Tan B P, et al. Effects of dietary administration of probiotic Halomonas sp. B12 on the intestinal micro?ora, immunological parameters, and midgut histological structure of shrimp, Fenneropenaeus chinensis. J. World Aquacult. Soc. 2009, 40: 58-66
Zhang Q, Ma H M, Mai K S, et al. Interaction of dietary Bacillus subtilis and fructooligosaccharide on the growth performance, non-specific immunity of sea cucumber, Apostichopus japonicus. Fish Shellfish Immunol., 2010, 29: 204-211
Zhang Q, Tan B P, Mai K S, et al., Dietary administration of Bacillus (B. licheniformis and B. subtilis) and isomaltooligosaccharide in?uences the intestinal micro?ora, immunological parameters and resistance against Vibrio alginolyticus in shrimp, Penaeus japonicus (Decapoda: Penaeidae). Aquaculture Research, doi:10.1111/j.1365-2109.2010.02677.x, 2010, 1-10
Zhou J, Song X L, Huang J, et al. Effects of dietary supplementation of A3α-peptidoglycan on innate immune responses and defense activity of Japanese flounder(Paralichthys olivaceus). Aquaculture, 2006, 251: 172-181
Zhou Q C, Buentello J A, Gatlin D M III. Effects of dietary prebiotics on growth performance, immune response and intestinal morphology of red drum (Sciaenops ocellatus). Aquaculture, 2010, 309: 253-257
Zhou X Q, Kuang S Y, Tang L. Effects of oligomannose and fructose levels on growth and transport of carp. Fish Sci.,2002, 21(2): 13-14
Zhou X Q, Li Y L. The effects of Bio-Mos on intestinal micro?ora and immune function of juvenile Jian carp (Cyprinus carpio Var. Jian). In: Nutritional biotechnology in the feed and food industries: Proceedings of Alltech’s 20th annual symposium (Suppl. 1-Abstracts of posters presented), Lexington, KY, USA, 2004
Zhou Z, Ding Z, Lv H Y. Effects of dietary short-chain fructooligosaccharides on intestinal microflora, survival and growth performance of juvenile white shrimp Litopenaeus vannamei. J. World Aquacult. Soc., 2007, 38: 296-301
Ziaei-Nejad S, Rezaei M H, Takami G A, et al. The effect of Bacillus spp. bacteria used as probiotics on digestive enzyme activity, survival and growth in the Indian white shrimp Fenneropenaeus indicus. Aquaculture, 2006, 252: 516-524
曹煜成,李卓佳,杨莺莺,等.地衣芽孢杆菌De株对黄鳍鲷生长及其养殖池塘主要环境因子的影响.南方水产,2010,6(3):1-6
曾东.鲤益生菌的筛选、生物学特性及作用机理的研究. [博士学位论文].绵阳:四川农业大学,2009
常青,梁萌青,王家林等.壳聚糖对花鲈生长和非特异性免疫力的影响.海洋水产研究,2006,27(5):17-22
陈国福,宋晓玲,黄倢,等. A3α-肽聚糖对凡纳滨对虾生长、免疫机能和抗病毒感染的影响.高技术通讯,2005,15(8):100-106
陈继红,王成章,王彦华,等.多糖在动物生产中的应用.饲料博览,2006,3,9-11
陈云波,华雪铭,周洪琪,等.壳聚糖对异育银鲫生长及抗菌能力的影响.上海水产大学学报,2006,15(2):243-246
程池,杨梅,李金霞,等. Biolog微生物自动分析系统——细菌鉴定操作规程的研究.食品与发酵工业,2006,32(5):50-54
方波,养殖刺参腐皮综合征病原学及其感染源的研究. [硕士学位论文].青岛:中国海洋大学,2006
付传永.枯草芽孢杆菌对刺参腐皮综合病致病菌的拮抗作用以及对水质的影响. [硕士学位论文].青岛:中国海洋大学,2008
付天玺,许国焕,吴月嫦,等.凝结芽孢杆菌对奥尼罗非鱼消化酶活性、消化率及生长性能的影响.淡水渔业,2008,38(4):30-35
郝向举.中华绒螯蟹肠道益生芽孢杆菌的筛选、生长特性及其应用效果的研究. [硕士学位论文].苏州:苏州大学,2010
何四旺,许国焕,吴月嫦,等.低聚异麦芽糖和低聚果糖对罗非鱼生长和非特异性免疫的影响.中国饲料,2003,23:14-15
胡毅,谭北平,麦康森,等.饲料中益生菌对凡纳滨对虾生长、肠道菌群及部分免疫指标的影响.中国水产科学,2008,15(2):244-251
胡毅.凡纳滨对虾饲料配方优化及几种饲料添加剂的应用. [博士学位论文].青岛:中国海洋大学,2007
华雪铭,周洪琪,张冬青,等.多糖和益生菌对暗纹东方鲀免疫功能的调节.水产学报,2006,30(2):230-235
华雪铭,周洪琪,张宇峰,等.饲料中添加壳聚糖和益生菌对暗纹东方鲀幼鱼生长及部分消化酶活性的影响.水生生物学报,2005,29(3):299-305
黄俊文,林映才,冯定远,等.益生菌、甘露寡糖对早期断奶仔猪生长、免疫和抗氧化机能的影响.动物营养学报,2005,17(4):16-20
黄美珍.光合细菌对致病弧菌的抑制作用.台湾海峡,1999,18(1):92-94
季高华,刘至治,冷向军.饲料中添加β-葡聚糖和低聚果糖对中华鳖幼鳖生长和血清SOD、溶菌酶活力的影响.上海水产大学学报,2004,13:36-40
李海兵,宋晓玲,韦嵩,等. 4株对虾肠道益生菌的筛选及鉴定.海洋与湖沼,2008,39,4:374-380
李海芳,常亚青,丁君.水产动物免疫增强剂的研究现状及应用前景.水产科学,2004,23(10):35-39
李运,盛慧,赵荣华.微生物鉴定系统在菌种鉴定中的应用.酿酒科技,2005,7:84-85
廖玉麟.我国的海参.生物学通报,2001,36(9):1-3
廖玉麟.中国动物志棘皮动物门海参纲.北京:科学出版社,1997. 53-64
刘君,宋晓玲,陈志鑫.益生菌对水产动物的作用研究进展.动物医学进展,2009,30(9):78-81
刘晓云,谭金山,包振民,等.刺参体腔细胞超微结构的观察.电子显微学报,2005,24(6):613-615
刘云,孔伟丽,姜国良,等. 2种免疫多糖对刺参组织主要免疫酶活性的影响.中国水产科学,2008,15(5):787-793
骆艺文.刺参有益菌制剂的研制与应用研究. [硕士学位论文].青岛:中国海洋大学,2009
马悦欣,徐高蓉,常亚青,等.大连地区刺参幼参溃烂病细菌性病原的初步研究.大连水产学院学报,2006a,21(1):13-18
马悦欣,徐高蓉,张恩鹏,等.仿刺参幼参急性口围肿胀症的细菌性病原.水产学报,2006b,30(3):377-382
麦康森,孟繁伊,马洪明,等.在仿刺参体腔液中发现新的调理素样分子.中国工程科学,2009,11(10): 108-114
孟凡伦,马桂荣,孔健.乳链球菌SB900肽聚糖对中国对虾免疫功能的影响.山东大学学报(自然科学版)1999a,34(1):88-93
孟凡伦,张玉臻,孔健,等.甲壳动物中的酚氧化酶原激活系统研究评价.海洋与湖沼,1999b,30(1):110-116
闵钟熳,牛天贵,岳喜庆.芽孢杆菌益生菌株的筛选.沈阳农业大学学报,2007,38(2):190-193
沈萍,范秀容,李广斌.微生物学实验第三版.北京:高等教育出版社,1999. 74-77
宋晓玲,王秀华,陈国福,等.肽聚糖制剂提高凡纳对虾抗白斑综合征病毒感染力的研究.高技术通讯,2005,15(1):74-78
孙永欣,王吉桥,汪婷婷,等.海参防御机制的研究进展.水产科学,2007,26(6):354-361
谭北平,周歧存,郑石轩,等.β-1,3/1,6-葡聚糖制剂对凡纳对虾生长及免疫力的影响.高技术通讯,2004,5:73-77
王斌,赵文,范薇,等.复方中药制剂对草鱼免疫细胞功能及抗病力影响的初步研究.大连水产学院学报,2007,22(3):203-206
王福强.牙鲆肠道益生菌的分离鉴定及其应用研究. [博士学位论文].北京:中国农业大学,2004
王高学,原居林,赵云奎,等.刺参表皮溃烂病病原菌的分离鉴定与药敏试验.西北农林科技大学学报(自然科学版),2007,35(8):87-91
王开来.维生素C对刺参生长和非特异性免疫的影响. [硕士学位论文].大连:大连理工大学,2009
王路平,吴垠,班红琴,等.微生态制剂对刺参幼参在封闭式循环养殖系统中的应用研究.中国微生态学杂志,2009,21(6):497-504
王秀华,黄倢,宋晓玲.免疫增强剂―肽聚糖在对虾养殖中的应用.海洋水产研究,2003,24(1):69-74
王秀华,宋晓玲,黄倢.肽聚糖制剂对南美白对虾体液免疫因子的影响.中国水产科学,2004b,11(1):26-30
王秀华,宋晓玲,黄倢.肽聚糖制剂对日本对虾非特异免疫因子的作用.高技术通讯,2004a,5:78-81
王印庚,方波,张春云,等.养殖刺参保苗期重大疾病“腐皮综合征”病原及其感染源分析.中国水产科学, 2006,13(4):610-616
王长法,张士璀,王昌留.水生无脊椎动物凝集素研究进展概述.海洋科学,2005,29(4):63-67
王正丽.免疫增强剂对牙鲆(Paralichthys Olivaceu)免疫力和抗病力的影响:[博士学位论文]. 青岛:中国海洋大学,2004
吴国忠,房如森,王松刚,等.壳聚糖的研究进展及其在水产养殖中的应用.中国水产, 2005,12,62-63
熊川男,李伟,白雪芳,等.凝集素作为海参免疫增强剂在人工养殖海参中的应用.饲料工业,2005,26(18):30-32
许乐乐.刺参(Apostichopus japonicas)体腔细胞原代培养技术的建立及其在快速筛选免疫增强剂中的应用. [博士学位论文].青岛:中国海洋大学,2009
闫大伟,华雪铭,周洪琪.壳聚糖对草鱼生长、抗病性能的影响.饲料工业,2007,28(12):17-18
阳会军,谭北平,方怀义.饲料中添加不同水平β-葡聚糖对斑节对虾生长、存活及抗病力的影响.饲料工业,2001,22(9):18-19
杨正梅,卜友泉,何瑞国.低聚果糖的生物学效应及其安全性研究进展.生命科学研究,2004,8(4):122-126
袁成玉,张洪,吴垠,等.微生态制剂对幼刺参生长及消化酶活性的影响.水产科学,2006,25(12):612-615
袁金凤.鸡源益生芽孢杆菌的筛选、发酵工艺优化及益生特性. [硕士学位论文].武汉:湖北工业大学,2009
张春晓,麦康森,艾庆辉,等.饲料中添加肽聚糖对大黄鱼生长和非特异性免疫力的影响. 水产学报,2008,32(3):411-416
张春云,王印庚,荣小军.养殖刺参腐皮综合征病原菌的分离与鉴定.水产学报,2006,30(1):118-123
张峰,宫晶,王海峰,等.仿刺参补体类似物活性的测定.大连水产学院学报,2007,22(4):246-248
张峰.棘皮动物体内防御机制研究进展.大连水产学院学报,2005,20(4):340-344
张玲,谭北平,麦康森,等.中国对虾体内1株益生菌的筛选与初步鉴定.中国海洋大学学报,2008,38(2):225-231
张玲.一株对虾肠道益生菌的筛选其作用机理和应用效果的研究. [博士学位论文].青岛:中国海洋大学,2007
张璐,艾庆辉,麦康森,等.肽聚糖对鲈鱼生长和非特异性免疫力的影响.中国海洋大学学报,2008,38(4):551-556
张琴.刺参高效免疫增强剂的筛选与应用. [博士学位论文].青岛:中国海洋大学,2010
张涛,白岚,李蕾,等.不同添加量的益生菌组合对仿刺参消化和免疫指标的影响.大连水产学院学报,2009,24(S):64-68
赵友福,魏亚东,高崇省,等.利用Biolog鉴定系统快速鉴定菜豆萎蔫病菌的研究. 1997,27(2):139-144
周慧慧,马洪明,张文兵,等.仿刺参肠道潜在益生菌对稚参生长、免疫及抗病力的影响.水产学报,2010,34(6):955-963
周慧慧.刺参(APoschopus jaPonicus)肠道益生菌的应用研究. [硕士学位论文].青岛:中国海洋大学,2010
朱伟,麦康森,张百刚,等.刺参稚参对蛋白质和脂肪需求量的初步研究.海洋科学,2005,29(3):54-58
Ainsworth A J. A beta-glucan inhibitable zymosan receptor on channel catfish neutrophils. Vet. Immunol Immunopathol., 1994, 41: 141-152
Aly S M, Abd-El-Rahman A M, John G, et al. Characterization of Some Bacteria Isolated from Oreochromis niloticus and their Potential Use as Probiotics. Aquaculture, 2008a, 277: 1-6
Aly S M, Ahmed Y A, Ghareeb A A, et al. Studies on Bacillus subtilis and Lactobacillus acidophilus, as potential probiotics, on the immune response and resistance of Tilapia nilotica (Oreochromis niloticus) to challenge infections. Fish Shellfish Immunol., 2008b, 25: 128-136
Aly S M, Mohamed M F, John G. Effect of probiotics on the survival, growth and challenge infection in Tilapia nilotica (Oreochromis niloticus). Aquaculture Research 2008c, 39: 647-656
Amann R I, Ludwig W, Schleifer K H. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev.,1995, 59: 143-169
Anderson D P, Siwicki A K, Rumsey G L. Injection or immersion delivery of selected immunostimulants to trout demonstrate enhancement of nonspecific defense mechanisms and protective immunity. Diseases in Asian Aquaculture, Vol.Ⅱ. Fish Health Section, Manila: Asia Fisheries Society, 1995, 413-426
Anderson D P, Siwicki A K. Duration of protection against Aeromonas salmonicida in brook trout immunostimulated with glucan or chitosan by injection or immersion. Pro. Fish Cult., 1994, 56: 258-261
Anderson D P. Immunostimulants, adjuvants, and vaccine carriers in fish: application to aquaculture. Fish Diseases, 1992(2): 281-307
Apún-Molina J P, Santamaría-Miranda A, Luna-Gonzalez A, et al. Effect of potential probiotic bacteria on growth and survival of tilapia Oreochromis niloticus L., cultured in the laboratory under high density and suboptimum temperature. Aquaculture Research, 2009, 40: 887- 894
Bagheri T, Hedayati S A, Yavari V, et al. Growth, Survival and Gut Microbial Load of Rainbow Trout (Onchorhynchus mykiss) Fry Given Diet Supplemented with Probiotic during the Two Months of First Feeding. Turkish Journal of Fisheries and Aquatic Sciences, 2008, 8: 43-48
Bagni M, Romano N, Finoia M G,et al. Short- and long-term effects of a dietary yeastβ-glucan (Macrogard) and alginic acid (Ergosan) preparation on immune response in sea bass (Dicentrarchus labrax). Fish Shellfish Immunol., 2005, 18: 311-325
Balcázar J L, de Blas I, Ruiz-Zarzuela I, et al. The role of probiotics in aquaculture. Veterinary Microbiology, 2006, 114: 173-186
Balcázar J L, Rojas-Luna T, Cunningham D P. E147-150
Balcázar J L. Evaluation of probiotic bacterial strains in Litopenaeus vannamei. Final Report, National Center for Marine and Aquaculture Research, Guayaquil, Ecuador. 2003
Bandyopadhyay P, Mohapatra P K D. Effect of a probiotic bacterium Bacillus circulans PB7 in the formulated diets: on growth, nutritional quality and immunity of Catla catla (Ham.) Fish Physiol. Biochem., 2009, 35: 467-478
Barracco M A, Duvic B, Soderhall K. Theβ-1,3-glucan binding protein from the crayfish Pacifastacus leniusculus, when reacted with aβ-1,3-glucan induces spreading and degranulation of crayfish granular cell. Cell Tissue Res, 1991, 266: 491-497
Bondad-Reantaso M, Subasinghe R P, Arthur J R, et al. Disease and health management in Asian aquaculture. Vet. Parasitology, 2005, 132: 249-272
Boolootian R A., Giese A C. Clotting of echinoderm coelomic fluid. J. Exper. Zool., 1959, 140:207-211
Boonyaratpalin S, Boonyyaratpalin M, Supamattaya K, et al. Effects of peptidoglycan (PG) on growth, survival, immune response and tolerance to stress in black tiger shrimp, Penaeus monodon. In: Diseases in Asian Aquaculture. Vol. 11. Fish Health Section, Asian Fisheries Society, Manila: Philippines, 1995, 469-477
Bornet F R, Brouns F. Immune-stimulating and gut health-promoting properties of short chain fructo-oligosaccharides. Nutr. Rev., 2002, 60(11): 326-334
Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Analytical Biochemistry, 1976, 72: 248-254
Brown G D, Gordon S. Fungalβ-Glucans and Mammalian Immunity. Immunity, 2003, 19, 311-315.
Buentello J A, Neill W H, Gatlin D M III, Effects of dietary prebiotics on the growth, feed efficiency and non-specific immunity of juvenile red drum Sciaenops ocellatus fed soybean-based diets. Aquaculture Research, 2010,41: 411-418
Bulgakov A A, Eliseikina M G, Petrova I Y, et al. Molecular and biological characterization of a mannan-binding lectin from the holothurian Apostichopus japonicus. Glycobiology,2007, 17: 1284-1288
Byun J W, Park S C, Benno Y, et al. Probiotic effect of Lactobacillus sp. DS-12 in ?ounder (Paralichthys olivaceus). J. Gen. Appl. Microbiol., 1997, 43: 305–308
Campa-Córdova A I, Hernández-Saavedra N Y, De Philippis R, et al. Generation of superoxide anion and SOD activity in haemocytes and muscle of American white shrimp(Litopenaeus vannamei) as a response toβ-glucan and sulphated polysaccharide. Fish Shellfish Immunol., 2002, 12: 353-366
Canicatti C, D'Ancona G, Cellular aspects of Holothuria polii immune response. J. Invertebr. Pathol., 1989, 53: 152-158
Canicatti C, G(o|¨)tz P. Dopa oxidation by Holothuria polii coelomocyte lysate. J. Invertebrate Pathol., 1991, 58: 305-310
Canicatti C, Lysosomal enzyme pattern in Holothuria polii coelomocytes. J. Invert Pathol., 1990, 56: 70-74
Canicatti C, Pagliara P, Stabili L. Sea urchin coelomic fluid agglutinin mediates coelomocyte adhesion. Eur. J. Cell Biol., 1992, 58(2): 291-295
Canicatti C, Parrinello N. Hemaglutinin and hemolysin level in coelomic fluid from Holothuria polii (Echinodermata) following sheep erythrocyte injection. Biol. Bull., 1985, 168: 175-182
Canicatti C, Rizzo A. A 220 kDa coelomocyte aggregating factor involved in Holothuria polii celluar clotting. Eur. J. cell Biol., 1991, 56: 79-83
Canicatti C, Seymour J. Evidence for phenoloxidase activity in Holothuria tubulosa (Echinodermata) brown bodies and cells. Parasitol. Res., 1991, 77: 50-53
Canicatti C. Hemolysins: pore-forming proteins in invertebrates. Celluar and Molecular life science, 1990, 46(3): 239-244
Cao Q Z, Lin Z B. Antitumor and anti-angiogenic activity of Ganoderma lucidum polysaccharides peptide. Acta Pharmacol. Sin., 2004, 25: 833-838
Castex M, Lemaire P, Wabete N, et al. Effect of probiotic Pediococcus acidilactici on antioxidant defences and oxidative stress of Litopenaeus stylirostris under Vibrio nigripulchritudo challenge. Fish Shellfish Immunol., 2010, 28: 622-631
Chang C F, Chen H Y, Su M S,et al. Immunomodulation by dietaryβ-1,3-glucan in the brooders of the black tiger shrimp Penaeus monodon Fish Shellfish Immunolo., 2000, 10: 505-514
Chang C F, Su M S, Chen H Y, et al. Dietaryβ-1,3-glucan effectively improves immunity and survival of Penaeus monodon challenged with white spot syndrome virus. Fish Shellfish Immunol., 2003, 15: 297-310
Chang Y Q, Yu C Q, Song X. Pond culture of sea cucumbers, Apostichopus japonicus, in Dalian. In: Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy, 2004. 269-272
Chen J X. Present status and prospects of sea cucumber industry in China. In: Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy, 2004. 25-38
Chia F S, Xing J. Echinoderm coelomocytes, a review. Zool. Stud., 1996, 35(4): 231-254 Chythanya R, Karunasager I, Karunasager I. Inhibition of shrimp pathogenic vibirios by a marine PseudomonasⅠ-2 strain. Aquaculture, 2002, 208: 1-10
Coffaro K A, Hinegardner A T. Immune response in the sea urchin Lytechinus oictus Science, 1977, 197: 1389-1390
Cook M T, Hayball P J, Hutchinson W, et al. Administration of a commercial immunostimulant preparation, EcoActiva as a feed supplement enhances macrophase respiratory burst and the growth rate of snapper (Pagrus auratus, Sparidae (Bloch and Schneider)) in winter. FishShellfish Immunol., 2003, 14: 333-345
Coteur G, Warnau M, Jangoux M, Dubois P. Reactive oxygen species (ROS) production by amoebocytes of Asterias rubens (Echinodermata). Fish Shellfish Immunol., 2002, 12(3): 187-200
Couso N, Castro R, Magarinos B, et al. Effect of oral administration of glucans on the resistance of gilthead seabream to pasteurellosis. Aquaculture, 2003, 219: 99-109
Culjak V, Bogut G, Has-Schon E, Milakovic Z, Canecki K (2006) Effect of Bio-Mos on performance and health of juvenile carp. In: Nutrition and biotechnology in the feed and food industries: Alltech’s 22nd annual symposium (suppl. 1-abstracts of posters presented), Lexington, KY, USA, 2006
Dalmin G, Kathiresan K, Purushothaman A. Effect of probiotics on bacterial population and health status of shrimp in culture pond ecosystem. Indian J. Exp. Biol., 2001, 39: 939-942
Dalmo R A, B(?)gwald J.β-glucans as conductors of immune symphonies. Fish Shellfish Immunol., 2008,25: 384-396
Daniels C L, Merrifield D L, Boothroyd D P, et al. Effect of dietary Bacillus spp. and mannan oligosaccharides (MOS) on European lobster (Homarus gammarus L.) larvae growth performance, gut morphology and gut microbiota. Aquaculture, 2010, 304: 49-57
Deeseenthum S, Leelavatcharamas V, Brooks J D. Effect of feeding Bacillus sp. As probiotic bacteria on growth of giant freshwater prawn (Macrobrachium rosenbergii de Man). Pakistan Journal of Biological Sciences, 2007, 10(9): 1481-1485
Delzenne N, Aertssens J, Verplaetse H, et al. Effect of fermentable fructo-oligosaccharides on mineral, nitrogen and energy digestive balance in the rat. Life Sci., 1995, 57: 1579-1587
Didier J, Philippe D., Michel J. Defensive mechanisms of holothriod(Echinodermata): Formation, role and fate of intracoelomic brown bodies in sea cucumber Holothuria tubulosa. Cell and Tissue Research, 1996, 283(1): 99-106
Dolmatova L S, Eliseikina M G, Romashina V V. Antioxidant enzymatic activity of coelomocytes of the far East sea cucumber Eupentacta fraudatrix. J. Evol. Biochem. Physiol., 2004, 40(2): 126-135
Du X J, Zhao X F, Wang J X. Molecular cloning and characterization of a lipopolysaccharide and β-1,3-glucan binding protein from fleshy prawn (Fenneropenaeus chinensis). Molecular Immunology 2007, 44: 1085-1094
Dybas L, Fankboner P V. Holothurian survival strategies: mechanisms for the maintenance of a bacteriostatic environment in the coelomic cavity of the sea cucumber, Parastichopus californicus. Dev. Comp. Immunol., 1986, 10(3): 311-330
El-Dakar A Y, Shalaby S M, Saoud I P. Assessing the use of a dietary probiotic/prebiotic as an enhancer of spinefoot rabbitfish Siganus rivulatus survival and growth. Aquaculture Nutrition, 2007, 13: 407-412
Eliseikina M G, Magarlamov T Y. Coelomocyte morphology in the holothurians Apostichopusjaponicus and Cucumaria japonica. Russian Journal of Marine Biology, 2002, 28(3): 197-202
El-Rhman A M A, Khattab Y A E, Shalaby A M E. Micrococcus luteus and Pseudomonas species as probiotics for promoting the growth performance and health of Nile tilapia, Oreochromis niloticus. Fish Shellfish Immunol., 2009, 27: 175-180
Engstad R E, Robertsen B. Specificity of aβ-glucan receptor on macrophages from Atlantic salmon (Salmo salar L.). Dev. Comp. Immunol., 1994, 18: 397-408
FAO/WHO. Joint FAO/WHO (Food and Agriculture Organization/World Health Organization) working group report on drafting guidelines for the evaluation of probiotics in food. London,
Ontario, Canada. guidelines for the evaluation of probiotics in food. Joint working group report on drafting. London, Ontario, 2002, pp. 1-11.
Farzanfar A. The use of probiotics in shrimp aquaculture. FEMS Immunol Med Microbiol. 2006, 48: 149-158
Fontaine A R., Lambert P. The fine structure of the leucocytes of the holothurian, Cucumaria miniata. Can. J. Zool., 1977, 55: 1530-1544
Franchini A, Conte A, Ottaviani E. Nitric oxide: an ancestral immunocyte effector molecule. Advances in Neuroimmunology, 1995, 5: 463-478
Fu L L, Shuai J B, Xu Z R, et al. Immune responses of Fenneropenaeus chinensis against white spot syndrome virus after oral delivery of VP28 using Bacillus subtilis as vehicles. Fish Shellfish Immunol., 2010, 28: 49-55
Fuller R. Probiotic in man and animals. J. Appl. Bacteriol., 1989, 66: 365-378
Gatesoupe F J. The use of probiotics in aquaculture, Aquaculture, 1999, 180: 147-165
Ghosh K, Sen S K, Ray A K. Supplementation of an isolated fish gut bacterium, Bacillus circulans, in formulated diets for rohu, Labeo rohita, fingerlings. Israeli J. Aquacult. Bamidgeh, 2003, 55: 13-21
Gibson G R, Roberfroid M B. Dietary modulation of the human colonic microbiota: introducing the concept of prebiotics. J. Nutr., 1995, 125: 1401-1412
Gildberg A, Mikkelsen H, Sandaker E, et al. Probiotic effect of lactic acid bacteria in the feed on growth and survival of fry of Atlantic cod (Gadus morhua). Hydrobiologia 1997, 352: 279-285
Goldin B R. Health benefits of probiotics. Br. J. Nutr., 1998, 80: S203-207
Gomez-Gil B, Roque A, Turnbull J F. The use and selection of probiotic bacteria for use in the culture of larval aquatic organisms. Aquaculture, 2000, 191: 259-270
Gopalakannan A, Arul V. Immunomodulatory effects of dietary intake of chitin, chitosan and levamisole on the immune system of Cyprinus carpio and control of Aeromonas hydrophila infection in ponds. Aquaculture, 2006, 255: 179-187
Gowda N M, Goswami U, Khan M I. Purification and characterization of a T-antigen specific lectin from the coelomic fluid of a marine invertebrate, sea cucumber (Holothuria scabra).Fish Shellfish Immunol., 2008, 24(4):450-458
Gram L, Melchiorsen J, Spanggaard B, et al. Inhibition of Vibrio anguillarum by Pseudomonasfluorescens strain AH2, a possible probiotic treatment of fish. Appl. Environ. Microbiol.,1999, 65, 969-973
Grisdale-Helland B, Helland S J, Gatlin D M III. The e ects of dietary supplementation withmannanoligosaccharide, fructooligosaccharide or galsctooligosaccharide on the growth andfeed utilization of Atlantic salmon (Salmo salar L.). Aquaculture, 2008, 283: 163-167
Gu M, Ma H M, Mai K S, Zhang W B, Ai Q H, Wang X J, Bai N. Immune response of seacucumber Apostichopus japonicus coelomocytes to several immunostimulants in vitro.Aquaculture, 2010, 306: 49-56
Gullian M, Thompson F, Rodriguez J. Selection of probiotic bacteria and study of theirimmunostimulatory effect in Penaeus vannamei. Aquaculture, 2004, 233: 1-14
Hai N V, Buller N, Fotedar R. Effects of probiotics (Pseudomonas synxantha and Pseudomonasaeruginosa) on the growth, survival and immune parameters of juvenile western king prawns(Penaeus latisulcatus Kishinouye, 1896). Aquaculture Research, 2009, 40: 590-602
Hatakeyama T, Nagatomo H, Yamasaki N. Interaction of the hemolytic lectin CEL-III from themarine invertebrate Cucumaria echinata with the erythrocyte membrane. J. Biol. Chem.,1995, 270(8): 3560-3564.
Hatakeyama T, Sato T, Taira E, et al. Characterization of the interaction of hemocytic lectin CEL-Ⅲfrom the marine invertebrate, cucumaria echinata,with artificial lipid membranes:involvement of neutral sphingoglycolipids in the pore-forming process. J Biochem(Tokyo),1999, 125(2): 277-284
Haug T, Kjuul A K, Styrvold O B, et al. Antibacterial activity in Strongylocentrotus droebachiensis(Echinoidea), Cucumaria frondosa (Holothuroidea), and Asterias rubens (Asteroidea). J.Invertebr. Pathol., 2002, 81(2):94-102
Hernández-López J, Gollas-Galvan T, Vargas-Albores F. Activation of the prophenoloxidasesystem of brown shrimp (Penaues californiensis Holmes) Comp. Biochem. Physiol., 1996,Part C, 113: 61-66
Hjelm M, Bergh ?, Riaza A, et al. Selection and identification of autochthonous potentialprobiotic bacteria from turbot larvae (Scophthalmus maximus) rearing units. Syst. Appl.Microbiol., 2004, 27: 360-371
Hua X M, Zhou H Q, Zhang D Q, et al. Effect of dietary chitosan and probiotics on diseaseresistance and immunity of obscure puffer (Fugu obscures) Journal of fisheries of China,2007, 31(4): 478-486
Huang C C, Song Y L. Maternal transmission of immunity to white spot syndrome associatedvirus (WSSV) in shrimp (Panaeus monodon). Dev. Comp. Immunol, 1999, 23: 545-552
Ibrahema M D, Fathi M, Mesalhy S, et al. Effect of dietary supplementation of inulin and vitaminC on the growth, hematology, innate immunity, and resistance of Nile tilapia (Oreochromisniloticus). Fish Shellfish Immunol., 2010, 29: 241-246
Isaeva V V, Korenbaun E S. Defense functions of coelomocytes and immunity of echinoderms. Soviet Journal of Marine Biology, 1990, (15): 353-363
Itami T, Asano M, Tokushige K, et al. Enhancement of disease resistance of kuruma shrimp, Penaeus japonicus, after oral administration of peptidoglycan derived from Bifidobacterium thermophilum. Aquaculture, 1998, 164: 277-288
Itami T, Kondo M, Uozu M, et al. Enhancement of resistance against Enterococcus seriolicida infection in yellowtail, Seriola quinqueradiata (Temninck and Schlegel), by oral administration of peptidoglycan derived from Bifidobacterium thermophilum. J Fish Dis, 1996, 19: 185-187
Jayaraj S S, Thiagarajan R, Arumugam M, et al. Isolation, purification and characterization of β-1,3-glucan binding protein from the plasma of marine mussel Perna viridis. Fish Shellfish Immunol., 2008, 24, 715-725
Jonathan P R, Smith L C, Mariano L C, et al. Genomic insights into the immune system of the sea urchin. Science, 2006, 314: 952-956
Karp R D, Hiloernann W H. Specific allograft reactivity in the sea star Dermasterias imbricate. Transplantation, 1976, 22: 434-439
Kesarcodi-Watson A, Kaspar H, Lategan M J, et al. Probiotics in aquaculture: The need, principles and mechanisms of action and screening processes. Aquaculture, 2008, 274: 1-14
Kodama H, Hirota Y, Mukamoto M, et al. Activation of rainbow trout (Oncorhynchus mykiss) phagocytes by muramyl dipeptide. Dev. Comp. Immunol., 1993, 7: 129-140
Kono T, Ponpornpisit A, Sakai M. The analysis of expressed genes in head kidney of common carp Cyprinus carpio L. stimulated with peptidoglycan. Aquaculture, 2004, 235: 37-52
Kono T, Sakai M. The analysis of expressed genes in the kidney of Japanese flounder, Paralichthys olivaceus, injected with the immunostimulant peptidoglycan. Fish Shellfish Immunol., 2001, 11: 357-366
Kozasa M. Toyocerin (Bacillus toyoi) as growth promotor for animal feeding. Microbiologie Aliments Nutrition, 1986, 4(1): 121-135
Kumar B P, Sahu N P, Saharan N, et al. Effect of dietary source and level of chitin on growth and survival of post-larvae Macrobrachium rosenbergii. J. Appl. Ichthyol. 2006, 22: 363-368
Kumar R, Mukherjee S C, Ranjan R, et al. Enhanced innate immune parameters in Labeo rohita (Ham.) following oral administration of Bacillus subtilis. Fish Shellfish Immunol., 2008, 24: 168-172
Kwak J K, Park S W, Koo J G, et al. Enhancement of the non-specific defense activities in carp (Cyprinus carpio) and flounder (Paralichthys olivaces) by oral administration of Schizophyllan. Acta Biotechnol, 2003, 23(4): 359-371
Li J Q, Tan B P, Mai K S. Dietary probiotic Bacillus OJ and isomaltooligosaccharides in?uence the intestine microbial populations, immune responses and resistance to white spot syndromevirus in shrimp (Litopenaeus vannamei). Aquaculture, 2009, 291: 35-40
Li J Y, Sun X Q, Zheng F R, et al. Screen and effect analysis of immunostimulants for sea cucumber, Apostichopus japonicas. Chinese Journal of Oceanology and Limnology, 2009, 27(1): 80-84
Li P, Burr G S, Gatlin D M III, et al. Dietary Supplementation of Short-Chain Fructooligosaccharides In?uences Gastrointestinal Microbiota Composition and Immunity Characteristics of Pacific White Shrimp, Litopenaeus vannamei, Cultured in a Recirculating System. The Journal of Nutrition, 2007, 137: 2763-2768
Li P, Wen Q, Gatlin D M III. Dose-dependent in?uences of dietaryβ-1,3-glucan on innate immunity and disease resistance of hybrid striped bass Morone chrysops×Morone saxatilis. Aquaculture Research, 2009, 40: 1578-1584
Lilley D M, Stillwell R J. Probiotics: growth promoting factors produced by micro-organisms. Science, 1965, 147: 747-748
Lin Y C, Vaseeharan B, Chen J C. Identification and phylogenetic analysis on lipopolysaccharide andβ-1,3-glucan binding protein (LGBP) of kuruma shrimp Marsupenaeus japonicus. Developmental and Comparative Immunology 2008, 32: 1260-1269
Liu F S, Li F H, Dong B, et al. Molecular cloning and characterisation of a pattern recognition protein, lipopolysaccharide andβ-1,3-glucan binding protein (LGBP) from Chinese shrimp Fenneropenaeus chinensis. Mol. Biol. Rep. 2009, 36: 471-477
López N, Cuzon G, Gaxiola G, et al. Physiological, nutritional, and immunological role of dietary β-1,3-glucan and ascorbic acid 2-monophosphate in Litopenaeus vannamei juveniles. Aquaculture, 2003, 224, 223-243
Lv H Y, Zhou Z G, Rudeaux F, et al. Effects of dietary short chain fructooligosaccharides on intestinal microflora, mortality and growth performance of Oreochromis aureus×O. niloticus. Chin. Anim. Sci., 2007, 19(6): 691-697
Mahious A S, Gatesoupe F J, Hervi M, et al. Effects of dietary inulin and oligosaccharides as prebiotics for weaning turbot Psetta maxima (Linneuas, C. 1758). Aquacult. Int., 2006, 14: 219-229
Manning T S, Gibson G R. Prebiotics. Best Pract. Res. Clin Gastroenterol, 2004, 18: 287-298
Martin F P, Collen M C. Bias in template-to-product ratios in multitemplate PCR. Appl. Environ. Microbiol., 1998, 64: 3724-3730
Matsui T, Ozeli Y, Suzuli M, et al. Purification and characterization of two Ca2+ dependent lectins from coelomic plasma of sea cucumber, Stichopus japonicus. J. Biochem., 1994, 116(5): 1127-1133
Meng Z, Shao J, Xiang L. CPG oligodeoxynucleotides activate grass carp (Ctenopharyngodon idellus) macrophages. Dev. Comp. Immunol. 2003, 27(4): 313-321
Merrifield D L, Dimitroglou A, Bradley G., et al. Probiotic applications for rainbow trout (Oncorhynchus mykiss Walbaum) I. Effects on growth performance, feed utilization,intestinal microbiota and related health criteria. Aqua. Nutr., 2010, 16: 504-510
Misra C K, Das B K, Mukherjee S C, et al. Effect of long term administration of dietaryβ-glucan on immunity, growth and survival of Labeo rohita fingerlings. Aquaculture, 2006b, 255: 82-94
Misra C K, Das B K, Mukherjee S C, et al. Effect of multiple injections ofβ-glucan on non-specific immune response and disease resistance in Labeo rohita fingerlings. Fish Shellfish Immunol., 2006a, 20: 305-319
Nayak S K, Swain P, Mukherjee S C. Effect of dietary supplementation of probiotic and vitamin C on the immune response of Indian major carp, Labeo rohita (Ham.) Fish Shellfish Immunol. 2007, 23: 892-896
Nayak S K. Probiotics and immunity: a fish perspective. Fish Shellfish Immunol., 2010, 29: 2-14
Newaj-Fyzul A, Adesiyun A A, Mutani A. et al. Bacillus subtilis AB1 controls Aeromonas infection in rainbow trout (Oncorhynchus mykiss, Walbaum). Journal of Applied Microbiology, 2007, 103: 1699-1706
Olivier G, Evelyn T, Lallier R. Immunity to Aeromoas salmonicida in coho salmon (Oncorhynchus kisutch) induced by modified Freund’s complete adjuvant: its non-specific nature and the probable role of mocrophages in the phenomenon. Dev. Comp. Immunol., 1985, 9: 419-432
Olsson C, Westerdahl A, Conway P L, et al. Intestinal colonization potential of turbot (Scophthalmus maximus L.) and dab(Limanda limanda)associated bacteria with inhibition effects against vibrio anguillarum. Appl. Environ. Micro., 1992, 58(3): 551-556
Pan, X, Wu, T, Song Z, et al. Immune responses and enhanced disease resistance in Chinese drum, Miichthys miiuy (Basilewsky), after oral administration of live or dead cells of Clostridium butyrium CB2. Journal of Fish Diseases, 2008, 31: 679-686
Parker R B. Probiotics, the other half of the antibiotics story. Anim. Nutr. Health, 1974, 29: 4-8
Pedro G S, César A B, Francisco R, et al. Lipopolysaccharides induce intestinal serum amyloid A expression in the sea cucumber Holothuria glaberrima. Dev. Comp. Immunol., 2003, 27(2): 105-110.
Powell A, Rowley A F. The effect of dietary chitin supplementation on the survival and immune reactivity of the shore crab, Carcinus maenas. Comparative Biochemistry and Physiology, 2007, 147(A): 122-128
Qi Z Z, Zhang X H, Boon N, et al. Probiotics in aquaculture of China -current state, problems and prospect. Aquaculture, 2009, 290: 15-21
Ratcliffe N A, Rowley A F. Ivertebrate blood cells. New York:Academic press New York, 1980. 513-526
Rattanachai A, Hirono I, Ohira,T, et al. Peptidoglycan inducible expression of a serine proteinase homologue from kuruma shrimp (Marsupenaeus japonicus). Fish Shellfish Immunol., 2005, 18: 39-48
Rengpipat S, Phianphak W, Piyatiratitivorakul S, et al. Effects of a probiotic bacterium on black tiger shrimp Penaeus monodon survival and growth. Aquaculture, 1998, 167: 301-313
Rengpipat S, Rukpratanporn S, Piyatiratitivorakul S, et al. Immunity enhancement in black tiger shrimp (Penaeus monodon) by a probiont bacterium (Bacillus S11). Aquaculture, 2000, 191: 271-288
Ring(?) E, Olsen T (?), Gifstad R A, et al. Prebiotic in aquaculture: a review. Aquaculture Nutrition, 2010,16: 117-136
Roch P, Canicatti ., Sammarco S. Tetrameric structure of the active phenoloxidase evidence in the coelomocytes of the echinoderm Holothuria tubulosa. Comp. Biochem. Physiol., 1992, 102B: 349-355
Rodríguez-Ramos T, Carpio Y, Bolívar J, et al. An inducible nitric oxide synthase (NOS) is expressed in hemocytes of the spiny lobster Panulirus argus: Cloning, characterization and expression analysis. Fish Shellfish Immunol., 2010, 29: 469-479
Rodríguez-Ramos T, Espinosa E, Hernández-López J, et al. Effects of Echerichia coli lipopolysaccharides and dissolved ammonia on immune response in southern white shrimp Litopenaeus schmitti. Aquaculture, 2008, 274: 118-125
Sahoo P K, Das A, Mohanty S. et al. Dietaryβ-1, 3-glucan improves the immunity and disease resistance of freshwater prawn Macrobrachium rosenbergii challenged with Aeromonas hydrophila. Aquaculture. Research, 2008, 39, 1574-1578
Sahoo P K, Mukherjee S C. The effect of dietary immunomodulation upon E.tarda vaccination in healthy and immunocompromised Indian Major Carp (L.rohita). Fish Shellfish Immunol., 2002, 12(1): 1-16
Sakai M. Current research statue of fish immunostimulant. Aquaculture, 1999, 172: 63-92
Salinas I, Cuesta A, Esteban M A, et al. Dietary administration of Lactobacillus delbrueckii and Bacillus subtilis, single or combined, on gilthead seabream cellular innate immune responses. Fish Shellfish Immunol., 2005 19: 67-77
Sealey W M, Barrows F T, Hang A, et al. Evaluation of the ability of barley genotypes containing different amounts ofβ-glucan to alter growth and disease resistance of rainbow trout Oncorhynchus mykiss. Animal Feed Science and Technology, 2008, 141: 115-128
Selvaraj V, Sampath K, Sekar V. Administration of yeast glucan enhances survival and some non-specific and specific immune parameters in carp (Cyprinus carpio) infected with Aeromonas hydrophila Fish Shellfish Immunol., 2005, 19(4): 293-306
Sghir A, Chow J M, Mackie R I. Continuous culture selection of bifidobacteria and lactobacilli from human fecal samples using fructooligosaccharide as selective substrate. J. Appl. Microbiol., 1998, 85: 769-777
Sharifuzzaman S M, Austin B. In?uence of probiotic feeding duration on disease resistance and immune parameters in rainbow trout. Fish Shellfish Immunol., 2009, 27: 440-445
Sisak F. Stimulation of phagocytosis as assessed by luminol-enhanced chemiluminescence andresponse to salmonella challenge of poultry fed diets containing mannan oligosaccharides. In: Lyons T.P., Jacqus K.A.(Eds.). Biotechnology in the feed industry, proceedings of Alltech’s 10th annual symposium. Nottingham, UK: Nottingham University Press. 1995
Siwicki A K, Anderson D P , Rumsey GL. Dietary intake of immunostimulants by rainbow trout affects non-specific immunity and protection against furunculosis.Vet Immunopathol, 1994, 41: 125-139
Smith L C, Britten R J, Davidson E H. Sea urchin genes expressed in activated coelomocytes are identified by expressed sequence tags. Complement homologues and other putative immune response genes suggest immune system homology within the deuterostomes. Journal of Immunology. 1996, 156: 593-602
Smith L C, Britten R J, Davidson E H. SpCoel1: A sea urchin profilin gene expressed specifically in coelomocytes in response to injury. Molecular Biology of the Cell, 1992, 3: 403-414
Smith L C, Clow L A, Terwilliger D P. The ancestral complement system in sea urchins. Immunol. Rev. 2001, 180:16-34
Smith P, Davey S. Evidence for the competitive excluding of Aeromonas salmonicida from fish with stress inducible furunculosis by a fluorescent pseudomonad. J. of Fish Disease, 1993, 16: 521-524
Smith V J. The echinoderms. In: Rafcliffe N A, Rowley A F (eds). Invertebrate blood cells, London: Academic Press, 1981. 513-562
Sogaard H. Microbials for feed: Beyond lactic acid bacteria. Feed international, 1990, 11: 22-37
Son V M, Chang C C, Wu M C, et al. Dietary administration of the probiotic, Lactobacillus plantarum, enhanced the growth, innate immune responses, and disease resistance of the grouper Epinephelus coioides. Fish Shellfish Immunol., 2009, 26: 691-698
Song H L, Hsieh Y T. Immunostimulation of tiger shrimp (Penaeus monodon) hemocytes for generation of microbicidal substances: analysis of reactive oxygen species. Dev. Comp. Immunol, 1994, 18(3): 201-209
Song Y L, Yu C I, Lien T W, et al. Haemolymph parameters of Pacific white shrimp (Litopenaeus vannamei) infected with Taura syndrome virus. Fish Shellfish Immunol., 2003, 14: 317-331
Sritunyalucksana K, Lee S Y, Soderhall K A.β-1,3-glucan binding protein from the black tiger shrimp Penaeus monodon. Dev. Comp. Immunol., 2002, 26: 237-245
Sritunyalucksana K, Sithisarn P, Withayachumnarnkul B, et al. Activation of prophenoloxidase, agglutinin and antibacterial activity in haemolymph of the black tiger prawn, Penaeus monodon, by immunostimulants. Fish Shellfish Immunol., 1999, 9: 21-30
Staykov Y, Denev S, Spring P. In?uence of dietary mannan oligosaccharides (Bio-Mos) on growth rate and immune function of common carp (Cyprinus carpio L.). In: Howell B, Flos R (eds) Lessons from the past to optimise the future. European Aquaculture Society, Special Publication No 35, 2005, pp 431-432
Staykov Y, Spring P, Denev S, et al. Effect of a mannan oligosaccharide on the growth performanceand immune status of rainbow trout (Oncorhynchus mykiss). Aquacult. Int., 2007, 15: 153-161
Sugita H, Hirose Y, Matsuo N, et al. Production of the antibacterial substance by Bacillus sp.strain NM 12, an intestinal bacterium of Japanese coastal fish. Aquaculture, 1998, 165(3-4): 269-280
Sun H L, Liang M Q, Yan J P, et al. Nutrient requirements and growth of the sea cucumber, Apostichopus japonicus In: Lovatelli A. (Eds.), Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy. 2004. pp. 327-331
Sun Y Z, Yang H L, Ma R L, et al. Probiotic applications of two dominant gut Bacillus strains with antagonistic activity improved the growth performance and immune responses of grouper Epinephelus coioides. Fish Shellfish Immunol., 2010, 29: 803-809
Sun Y, Jin L, Wang T. Polysaccharides from Astragalus membranaceus promote phagocytosis and superoxide anion (O2-) production by coelomocytes from sea cucumber Apostichopus japonicus in vitro. Comp. Biochem. Physiol., 2008, 147(C):2 93-298
Suzer C, (?)oban D, , Kamaci H O, et al. Lactobacillus spp. bacteria as probiotics in gilthead sea bream (Sparus aurata, L.) larvae: Effects on growth performance and digestive enzyme activities. Aquaculture, 2008, 280: 140-145
Swain S M, Singh C, Arul V. Inhibitory activity of probiotics Streptococcus phocae PI80 and Enterococcus faecium MC13 against Vibriosis in shrimp Penaeus monodon. World J. Microbiol. Biotechnol., 2009, 25: 697-703
Sweetman J W, Torrecillas S, Dimitroglou A, et al. REVIEW ARTICLE Enhancing the natural defences and barrier protection of aquaculture species. Aquaculture Research, 2010, 41, 345-355
Taoka Y, Maeda H, Jo J Y, et al. Growth, stress tolerance and non-specific immune response of Japanese flounder Paralichthys olivaceus to probiotics in a closed recirculating system. Fish Sci., 2006, 72, 310-321
Tlaskalová-HogenováH, StepánkováR, Hudcovic T, et al. Commensal bacteria (normal microflora), mucosal immunity and chronic inflammatory and autoimmune diseases. Immunol. Lett., 2004, 93: 97-108
Torrecillas S, Makol A, Caballero M J, et al. Immune stimulation and improved infection resistance in European sea bass (Dicentrarchus labrax) fed mannan oligosaccharides. Fish Shellfish Immunol., 2007, 23: 969-981
Tseng D Y, Ho P L, Huang S Y, et al. Enhancement of immunity and disease resistance in the white shrimp, Litopenaeus vannamei, by the probiotic, Bacillus subtilis E20. Fish Shellfish Immunol., 2009, 26: 339-344
Vargas-Albores F, Yepiz-Plascencia G. Beta glucan binding protein and its role in shrimp immune response. Aquaculture, 2000, 191, 13-21
Vaseeharan B, Ramasamy P. Control of pathogenic Vibrio spp. by Bacillus subtilis BT23, apossible probiotic treatment for black tiger shrimp Penaeus monodon. Letters in applied microbiology, 2003, 36: 83-87
Vendrell D, Balcázar J L, de Blas, I. et al. Protection of rainbow trout (Oncorhynchus mykiss) from lactococcosis by probiotic bacteria. Comparative Immunology, Microbiology and Infectious Diseases, 2008, 31: 337-345
Venkat H K, Sahu N P, Jain K K. Effect of feeding Lactobacillus-based probiotics on the gut micro?ora, growth and survival of postlarvae of Macrobrachium rosenbergii (de Man). Aquaculture Research, 2004, 35: 501-507
Verschuere L, Rombaut G, Sorgeloos P, et al. Probiotic bacteria as biological control agents in aquaculture. Microbiology and Molecular Biology Review, 2000, 64: 655-671
Wang S H, Chen J C. The protective effect of chitin and chitosan against Vibrio alginolyticus in white shrimp Litopenaeus vannamei Fish Shellfish Immunol., 2005, 19: 191-204
Wang Y B, Han J Z. The role of probiotic cell wall hydrophobicity in bioremediation of aquaculture. Aquaculture, 2007, 269: 349-354
Wang Y B, Li J R, Lin J. Probiotics in aquaculture: Challenges and outlook Aquaculture, 2008, 281: 1-4
Wang Y B. Effect of probiotics on growth performance and digestive enzyme activity of the shrimp Penaeus vannamei. Aquaculture, 2007, 269: 259-264
Wang Y C, Chang P S, Chen H Y. Differential time-series expression of immune-related genes of Pacific white shrimp Litopenaeus vannamei in response to dietary inclusion of β-1,3-glucan.Fish Shellfish Immunol., 2008, 24: 113-121
Wang Y G, Zhang C Y, Rong X J, et al. Diseases of cultured sea cucumber, Apostichopus japonicus, in China. In: Lovatelli A. (Ed.), Advances in sea cucumber aquaculture and management. Food and Agriculture Organization of the United Nations. Rome, Italy. 2004, pp. 297-310
Wang Y J, Osatomi K, Yoshida A, et al. Extracellular products from virulent strain of Edwardsiella tarda stimulate mouse macrophages (RAW264.7) to produce nitric oxide (NO) and tumor necrosis factor (TNF)-α. Fish Shellfish Immunol., 2010, 29: 778-785
Whittington R, Lim C, Klesius P H. Effect of dietaryβ-glucan levels on the growth response and efficacy of Streptococcus iniae vaccine in Nile tilapia, Oreochromis niloticus. Aquaculture, 2005, 248: 217-225
Xing J, chia F S. Opsonin-like molecule found in coelomic fluid of sea cucumber, Holothuria leucospilota. Marine Biology, 2000, 136: 979-986
Xing J, Leung M F, Chia F S. Quantitative analysis of phagcytosis by amoebocytes in a sea cucumber Holothuria leucospilota. Invert. Biol., 1998, 117: 67-74
Yeh F C, Wu S H, Lai C Y, et al. Demonstration of nitric oxide synthase activity in crustacean hemocytes and anti-microbial activity of hemocyte-derived nitric oxide. Comparative Biochemistry and Physiology, 2006, Part B 144(1):11-17
Yousefian M, Amiri M S, A review of the use of prebiotic in aquaculture for fish and shrimp. African Journal of Biotechnology, 2009, 8(25): 7313-7318
Zhang L, Mai K S, Tan B P, et al. Effects of dietary administration of probiotic Halomonas sp. B12 on the intestinal micro?ora, immunological parameters, and midgut histological structure of shrimp, Fenneropenaeus chinensis. J. World Aquacult. Soc. 2009, 40: 58-66
Zhang Q, Ma H M, Mai K S, et al. Interaction of dietary Bacillus subtilis and fructooligosaccharide on the growth performance, non-specific immunity of sea cucumber, Apostichopus japonicus. Fish Shellfish Immunol., 2010, 29: 204-211
Zhang Q, Tan B P, Mai K S, et al., Dietary administration of Bacillus (B. licheniformis and B. subtilis) and isomaltooligosaccharide in?uences the intestinal micro?ora, immunological parameters and resistance against Vibrio alginolyticus in shrimp, Penaeus japonicus (Decapoda: Penaeidae). Aquaculture Research, doi:10.1111/j.1365-2109.2010.02677.x, 2010, 1-10
Zhou J, Song X L, Huang J, et al. Effects of dietary supplementation of A3α-peptidoglycan on innate immune responses and defense activity of Japanese flounder(Paralichthys olivaceus). Aquaculture, 2006, 251: 172-181
Zhou Q C, Buentello J A, Gatlin D M III. Effects of dietary prebiotics on growth performance, immune response and intestinal morphology of red drum (Sciaenops ocellatus). Aquaculture, 2010, 309: 253-257
Zhou X Q, Kuang S Y, Tang L. Effects of oligomannose and fructose levels on growth and transport of carp. Fish Sci.,2002, 21(2): 13-14
Zhou X Q, Li Y L. The effects of Bio-Mos on intestinal micro?ora and immune function of juvenile Jian carp (Cyprinus carpio Var. Jian). In: Nutritional biotechnology in the feed and food industries: Proceedings of Alltech’s 20th annual symposium (Suppl. 1-Abstracts of posters presented), Lexington, KY, USA, 2004
Zhou Z, Ding Z, Lv H Y. Effects of dietary short-chain fructooligosaccharides on intestinal microflora, survival and growth performance of juvenile white shrimp Litopenaeus vannamei. J. World Aquacult. Soc., 2007, 38: 296-301
Ziaei-Nejad S, Rezaei M H, Takami G A, et al. The effect of Bacillus spp. bacteria used as probiotics on digestive enzyme activity, survival and growth in the Indian white shrimp Fenneropenaeus indicus. Aquaculture, 2006, 252: 516-524
曹煜成,李卓佳,杨莺莺,等.地衣芽孢杆菌De株对黄鳍鲷生长及其养殖池塘主要环境因子的影响.南方水产,2010,6(3):1-6
曾东.鲤益生菌的筛选、生物学特性及作用机理的研究. [博士学位论文].绵阳:四川农业大学,2009
常青,梁萌青,王家林等.壳聚糖对花鲈生长和非特异性免疫力的影响.海洋水产研究,2006,27(5):17-22
陈国福,宋晓玲,黄倢,等. A3α-肽聚糖对凡纳滨对虾生长、免疫机能和抗病毒感染的影响.高技术通讯,2005,15(8):100-106
陈继红,王成章,王彦华,等.多糖在动物生产中的应用.饲料博览,2006,3,9-11
陈云波,华雪铭,周洪琪,等.壳聚糖对异育银鲫生长及抗菌能力的影响.上海水产大学学报,2006,15(2):243-246
程池,杨梅,李金霞,等. Biolog微生物自动分析系统——细菌鉴定操作规程的研究.食品与发酵工业,2006,32(5):50-54
方波,养殖刺参腐皮综合征病原学及其感染源的研究. [硕士学位论文].青岛:中国海洋大学,2006
付传永.枯草芽孢杆菌对刺参腐皮综合病致病菌的拮抗作用以及对水质的影响. [硕士学位论文].青岛:中国海洋大学,2008
付天玺,许国焕,吴月嫦,等.凝结芽孢杆菌对奥尼罗非鱼消化酶活性、消化率及生长性能的影响.淡水渔业,2008,38(4):30-35
郝向举.中华绒螯蟹肠道益生芽孢杆菌的筛选、生长特性及其应用效果的研究. [硕士学位论文].苏州:苏州大学,2010
何四旺,许国焕,吴月嫦,等.低聚异麦芽糖和低聚果糖对罗非鱼生长和非特异性免疫的影响.中国饲料,2003,23:14-15
胡毅,谭北平,麦康森,等.饲料中益生菌对凡纳滨对虾生长、肠道菌群及部分免疫指标的影响.中国水产科学,2008,15(2):244-251
胡毅.凡纳滨对虾饲料配方优化及几种饲料添加剂的应用. [博士学位论文].青岛:中国海洋大学,2007
华雪铭,周洪琪,张冬青,等.多糖和益生菌对暗纹东方鲀免疫功能的调节.水产学报,2006,30(2):230-235
华雪铭,周洪琪,张宇峰,等.饲料中添加壳聚糖和益生菌对暗纹东方鲀幼鱼生长及部分消化酶活性的影响.水生生物学报,2005,29(3):299-305
黄俊文,林映才,冯定远,等.益生菌、甘露寡糖对早期断奶仔猪生长、免疫和抗氧化机能的影响.动物营养学报,2005,17(4):16-20
黄美珍.光合细菌对致病弧菌的抑制作用.台湾海峡,1999,18(1):92-94
季高华,刘至治,冷向军.饲料中添加β-葡聚糖和低聚果糖对中华鳖幼鳖生长和血清SOD、溶菌酶活力的影响.上海水产大学学报,2004,13:36-40
李海兵,宋晓玲,韦嵩,等. 4株对虾肠道益生菌的筛选及鉴定.海洋与湖沼,2008,39,4:374-380
李海芳,常亚青,丁君.水产动物免疫增强剂的研究现状及应用前景.水产科学,2004,23(10):35-39
李运,盛慧,赵荣华.微生物鉴定系统在菌种鉴定中的应用.酿酒科技,2005,7:84-85
廖玉麟.我国的海参.生物学通报,2001,36(9):1-3
廖玉麟.中国动物志棘皮动物门海参纲.北京:科学出版社,1997. 53-64
刘君,宋晓玲,陈志鑫.益生菌对水产动物的作用研究进展.动物医学进展,2009,30(9):78-81
刘晓云,谭金山,包振民,等.刺参体腔细胞超微结构的观察.电子显微学报,2005,24(6):613-615
刘云,孔伟丽,姜国良,等. 2种免疫多糖对刺参组织主要免疫酶活性的影响.中国水产科学,2008,15(5):787-793
骆艺文.刺参有益菌制剂的研制与应用研究. [硕士学位论文].青岛:中国海洋大学,2009
马悦欣,徐高蓉,常亚青,等.大连地区刺参幼参溃烂病细菌性病原的初步研究.大连水产学院学报,2006a,21(1):13-18
马悦欣,徐高蓉,张恩鹏,等.仿刺参幼参急性口围肿胀症的细菌性病原.水产学报,2006b,30(3):377-382
麦康森,孟繁伊,马洪明,等.在仿刺参体腔液中发现新的调理素样分子.中国工程科学,2009,11(10): 108-114
孟凡伦,马桂荣,孔健.乳链球菌SB900肽聚糖对中国对虾免疫功能的影响.山东大学学报(自然科学版)1999a,34(1):88-93
孟凡伦,张玉臻,孔健,等.甲壳动物中的酚氧化酶原激活系统研究评价.海洋与湖沼,1999b,30(1):110-116
闵钟熳,牛天贵,岳喜庆.芽孢杆菌益生菌株的筛选.沈阳农业大学学报,2007,38(2):190-193
沈萍,范秀容,李广斌.微生物学实验第三版.北京:高等教育出版社,1999. 74-77
宋晓玲,王秀华,陈国福,等.肽聚糖制剂提高凡纳对虾抗白斑综合征病毒感染力的研究.高技术通讯,2005,15(1):74-78
孙永欣,王吉桥,汪婷婷,等.海参防御机制的研究进展.水产科学,2007,26(6):354-361
谭北平,周歧存,郑石轩,等.β-1,3/1,6-葡聚糖制剂对凡纳对虾生长及免疫力的影响.高技术通讯,2004,5:73-77
王斌,赵文,范薇,等.复方中药制剂对草鱼免疫细胞功能及抗病力影响的初步研究.大连水产学院学报,2007,22(3):203-206
王福强.牙鲆肠道益生菌的分离鉴定及其应用研究. [博士学位论文].北京:中国农业大学,2004
王高学,原居林,赵云奎,等.刺参表皮溃烂病病原菌的分离鉴定与药敏试验.西北农林科技大学学报(自然科学版),2007,35(8):87-91
王开来.维生素C对刺参生长和非特异性免疫的影响. [硕士学位论文].大连:大连理工大学,2009
王路平,吴垠,班红琴,等.微生态制剂对刺参幼参在封闭式循环养殖系统中的应用研究.中国微生态学杂志,2009,21(6):497-504
王秀华,黄倢,宋晓玲.免疫增强剂―肽聚糖在对虾养殖中的应用.海洋水产研究,2003,24(1):69-74
王秀华,宋晓玲,黄倢.肽聚糖制剂对南美白对虾体液免疫因子的影响.中国水产科学,2004b,11(1):26-30
王秀华,宋晓玲,黄倢.肽聚糖制剂对日本对虾非特异免疫因子的作用.高技术通讯,2004a,5:78-81
王印庚,方波,张春云,等.养殖刺参保苗期重大疾病“腐皮综合征”病原及其感染源分析.中国水产科学, 2006,13(4):610-616
王长法,张士璀,王昌留.水生无脊椎动物凝集素研究进展概述.海洋科学,2005,29(4):63-67
王正丽.免疫增强剂对牙鲆(Paralichthys Olivaceu)免疫力和抗病力的影响:[博士学位论文]. 青岛:中国海洋大学,2004
吴国忠,房如森,王松刚,等.壳聚糖的研究进展及其在水产养殖中的应用.中国水产, 2005,12,62-63
熊川男,李伟,白雪芳,等.凝集素作为海参免疫增强剂在人工养殖海参中的应用.饲料工业,2005,26(18):30-32
许乐乐.刺参(Apostichopus japonicas)体腔细胞原代培养技术的建立及其在快速筛选免疫增强剂中的应用. [博士学位论文].青岛:中国海洋大学,2009
闫大伟,华雪铭,周洪琪.壳聚糖对草鱼生长、抗病性能的影响.饲料工业,2007,28(12):17-18
阳会军,谭北平,方怀义.饲料中添加不同水平β-葡聚糖对斑节对虾生长、存活及抗病力的影响.饲料工业,2001,22(9):18-19
杨正梅,卜友泉,何瑞国.低聚果糖的生物学效应及其安全性研究进展.生命科学研究,2004,8(4):122-126
袁成玉,张洪,吴垠,等.微生态制剂对幼刺参生长及消化酶活性的影响.水产科学,2006,25(12):612-615
袁金凤.鸡源益生芽孢杆菌的筛选、发酵工艺优化及益生特性. [硕士学位论文].武汉:湖北工业大学,2009
张春晓,麦康森,艾庆辉,等.饲料中添加肽聚糖对大黄鱼生长和非特异性免疫力的影响. 水产学报,2008,32(3):411-416
张春云,王印庚,荣小军.养殖刺参腐皮综合征病原菌的分离与鉴定.水产学报,2006,30(1):118-123
张峰,宫晶,王海峰,等.仿刺参补体类似物活性的测定.大连水产学院学报,2007,22(4):246-248
张峰.棘皮动物体内防御机制研究进展.大连水产学院学报,2005,20(4):340-344
张玲,谭北平,麦康森,等.中国对虾体内1株益生菌的筛选与初步鉴定.中国海洋大学学报,2008,38(2):225-231
张玲.一株对虾肠道益生菌的筛选其作用机理和应用效果的研究. [博士学位论文].青岛:中国海洋大学,2007
张璐,艾庆辉,麦康森,等.肽聚糖对鲈鱼生长和非特异性免疫力的影响.中国海洋大学学报,2008,38(4):551-556
张琴.刺参高效免疫增强剂的筛选与应用. [博士学位论文].青岛:中国海洋大学,2010
张涛,白岚,李蕾,等.不同添加量的益生菌组合对仿刺参消化和免疫指标的影响.大连水产学院学报,2009,24(S):64-68
赵友福,魏亚东,高崇省,等.利用Biolog鉴定系统快速鉴定菜豆萎蔫病菌的研究. 1997,27(2):139-144
周慧慧,马洪明,张文兵,等.仿刺参肠道潜在益生菌对稚参生长、免疫及抗病力的影响.水产学报,2010,34(6):955-963
周慧慧.刺参(APoschopus jaPonicus)肠道益生菌的应用研究. [硕士学位论文].青岛:中国海洋大学,2010
朱伟,麦康森,张百刚,等.刺参稚参对蛋白质和脂肪需求量的初步研究.海洋科学,2005,29(3):54-58